Complement factor b (cfb) irna compositions and methods of use thereof

ABSTRACT

The present invention relates to RNAi agents, e.g., dsRNA agents, targeting the complement factor B (CFB) gene. The invention also relates to methods of using such RNAi agents to inhibit expression of a CFB gene and to methods of treating or preventing a CFB-associated disease in a subject.

RELATED APPLICATION

This application is a 35 § U.S.C. 111(a) continuation application whichclaims the benefit of priority to PCT/US2022/047987, filed on Oct. 27,2022, which, in turn, claims the benefit of priority to U.S. ProvisionalApplication No. 63/273,215, filed on Oct. 29, 2021. The entire contentsof each of the foregoing applications are incorporated herein byreference.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted electronically in XML file format and is hereby incorporatedby reference in its entirety. Said XML copy, created on Mar. 21, 2023,is named 121301-18902_SL.xml and is 20,880,720 bytes in size.

BACKGROUND OF THE INVENTION

Complement was first discovered in the 1890s when it was found to aid or“complement” the killing of bacteria by heat-stable antibodies presentin normal serum (Walport, M. J. (2001) N Engl J Med. 344:1058). Thecomplement system consists of more than 30 proteins that are eitherpresent as soluble proteins in the blood or are present asmembrane-associated proteins. Activation of complement leads to asequential cascade of enzymatic reactions, known as complementactivation pathways resulting in the formation of the potentanaphylatoxins C3a and C5a that elicit a plethora of physiologicalresponses that range from chemoattraction to apoptosis. Initially,complement was thought to play a major role in innate immunity where arobust and rapid response is mounted against invading pathogens.However, recently it is becoming increasingly evident that complementalso plays an important role in adaptive immunity involving T and Bcells that help in elimination of pathogens (Dunkelberger J R and Song WC. (2010) Cell Res. 20:34; Molina H, et al. (1996) Proc Natl Acad SciUSA. 93:3357), in maintaining immunologic memory preventing pathogenicre-invasion, and is involved in numerous human pathological states (Qu,H, et al. (2009) Mol Immunol. 47:185; Wagner, E. and Frank M M. (2010)Nat Rev Drug Discov. 9:43).

Complement activation is known to occur through three differentpathways: alternate, classical and lectin (FIG. 1 ) involving proteinsthat mostly exist as inactive zymogens that are then sequentiallycleaved and activated.

The classical pathway is often activated by antibody-antigen complexesor by the C-reactive protein (CRP), both of which interact withcomplement component C1q. In addition, the classical pathway can beactivated by phosphatidyl serine present in apoptotic bodies in theabsence of immune complexes.

The lectin pathway is initiated by the mannose-binding lectins (MBL)that bind to complex carbohydrate residues on the surface of pathogens.The activation of the classical pathway or the lectin pathway leads toactivation of the (C4b2b) C3 convertase.

The alternate pathway is activated by the binding of C3b, which isspontaneously generated by the hydrolysis of C3, on targeted surfaces.This surface-bound C3b is then recognized by factor B, forming thecomplex C3bB. The C3bB complex, in turn, is cleaved by factor D to yieldthe active form of the C3 convertase of the AP (C3bBb). Both types of C3convertases will cleave C3, forming C3b. C3b then either binds to morefactor B, enhancing the complement activation through the AP (theso-called alternative or amplification loop), or leads to the formationof the active C5 convertase (C3bBbC3b or C4bC2bC3b), which cleaves C5and triggers the late events that result in the formation of themembrane attack complex (MAC) (C5b-9).

Inappropriate activation of the complement system is responsible forpropagating or initiating pathology in many different diseases,including, for example, C3 glomerulopathy, systemic lupus erythematosus(SLE), e.g., Lupus Nephritis, IgA nephropathy, diabetic nephropathy,polycystic kidney disease, membranous nephropathy, age-related maculardegeneration, atypical hemolytic uremic syndrome, thromboticmicroangiopathy, myasthenia gravis, ischemia and reperfusion injury,paroxysmal nocturnal hemoglobinuria, and rheumatoid arthritis.

To date, only one therapeutic that targets the alternate pathway, e.g.,the C5-C5a axis, is available for the treatment of complementcomponent-associated diseases, the anti-C5 antibody, eculizumab(Soliris®). Although eculizumab has been shown to be effective for thetreatment of paroxysmal nocturnal hemoglobinuria (PNH), atypicalhemolytic uremic syndrome (aHUS), and Myasthenia Gravis, and iscurrently being evaluated in clinical trials for additional complementcomponent-associated diseases, eculizumab therapy requires weekly highdose infusions followed by biweekly maintenance infusions at a highcost. Furthermore, approximately 50% of eculizumab-treated PNH subjectshave low level of hemolysis and require residual transfusions (Hill A,et al. (2010) Haematologica 95(4):567-73).

Accordingly, there is a need in the art for compositions and methods fortreating diseases, disorders, and conditions associated with complementactivation by, for example, activation of complement factor B activity.

SUMMARY OF THE INVENTION

The present invention provides iRNA compositions which affect theRNA-induced silencing complex (RISC)-mediated cleavage of RNAtranscripts of a gene encoding complement factor B (CFB). The complementfactor B (CFB) may be within a cell, e.g., a cell within a subject, suchas a human subject.

Accordingly, in one aspect, the invention provides a double strandedribonucleic acid (dsRNA) agent for inhibiting expression of complementfactor B (CFB) in a cell, wherein the dsRNA agent comprises a sensestrand and an antisense strand forming a double stranded region, whereinthe sense strand comprises at least 15, e.g., 15, 16, 17, 18, 19, 20, or21, contiguous nucleotides differing by no more than 0, 1, 2, or 3nucleotides from the nucleotide sequence of SEQ ID NO:1 and theantisense strand comprises at least 15, e.g., 15, 16, 17, 18, 19, 20,21, 22, or 23, contiguous nucleotides differing by no more than 1, 2, or3 nucleotides from the corresponding portion of the nucleotide sequenceof SEQ ID NO:8.

In another aspect, the present invention provides a double strandedribonucleic acid (dsRNA) for inhibiting expression of complement factorB (CFB) in a cell, wherein said dsRNA comprises a sense strand and anantisense strand forming a double stranded region, wherein the antisensestrand comprises a region of complementarity to an mRNA encodingcomplement factor B (CFB), and wherein the region of complementaritycomprises at least 15, e.g., 15, 16, 17, 18, 19, 20, 21, 22, or 23,contiguous nucleotides differing by no more than 0, 1, 2, or 3nucleotides from any one of the antisense nucleotide sequences in anyone of Tables 2-3.

In one embodiment, the dsRNA agent comprises a sense strand comprisingany of the sense nucleotide sequences in any one of Tables 2-3.

In one embodiment, the dsRNA agent comprises an antisense strandcomprising any of the antisense nucleotide sequences in any one ofTables 2-3.

In one embodiment, the dsRNA agent comprises a sense strand comprisingany of the sense nucleotide sequences in any one of Tables 2-3 and anantisense strand comprising any of the antisense nucleotide sequences inany one of Tables 2-3.

In one embodiment, the dsRNA agent comprises a sense strand comprising anucleotide sequence which differs by no more than 4 nucleotides from anyof the sense nucleotide sequences in any one of Tables 2-3 and anantisense strand comprising a nucleotide sequence which differs by nomore than 4 nucleotides from any of the antisense nucleotide sequencesin any one of Tables 2-3.

In one embodiment, the dsRNA agent comprises a sense strand comprising anucleotide sequence which differs by no more than 3 nucleotides from anyof the sense nucleotide sequences in any one of Tables 2-3 and anantisense strand comprising a nucleotide sequence which differs by nomore than 3 nucleotides from any of the antisense nucleotide sequencesin any one of Tables 2-3.

In one embodiment, the dsRNA agent comprises a sense strand comprising anucleotide sequence which differs by no more than 2 nucleotides from anyof the sense nucleotide sequences in any one of Tables 2-3 and anantisense strand comprising a nucleotide sequence which differs by nomore than 2 nucleotides from any of the antisense nucleotide sequencesin any one of Tables 2-3.

In one embodiment, the dsRNA agent comprises a sense strand comprising anucleotide sequence which differs by no more than 1 nucleotide from anyof the sense nucleotide sequences in any one of Tables 2-3 and anantisense strand comprising a nucleotide sequence which differs by nomore than 1 nucleotide from any of the antisense nucleotide sequences inany one of Tables 2-3.

In another aspect, the present invention provides a double strandedribonucleic acid (dsRNA) for inhibiting expression of complement factorB (CFB) in a cell, wherein said dsRNA comprises a sense strand and anantisense strand forming a double stranded region, wherein the sensestrand comprises at least 15, e.g., 15, 16, 17, 18, 19, 20, or 21,contiguous nucleotides differing by no more than 0, 1, 2, 3 or 4nucleotides from any one of the nucleotide sequence of nucleotides504-526, 640-662, 641-663, 995-1017, 997-1019, 1034-1056, 1141-1163,1145-1167, 1389-1411, 1473-1495, 1826-1848, 1828-1850, 1842-1864,2242-2264, 2391-2413, 2393-2415, 2438-2460, or 2453-2475 of SEQ ID NO:1, and the antisense strand comprises at least 15, e.g., 15, 16, 17, 18,19, 20, 21, 22 or 23, contiguous nucleotides from the correspondingnucleotide sequence of SEQ ID NO:8.

In one embodiment, the antisense strand comprises at least 15, e.g., 15,16, 17, 18, 19, 20, 21, 22, or 23, contiguous nucleotides differing byno more than 0, 1, 2, 3, or 4 nucleotides from any one of the antisensestrand nucleotide sequences of a duplex selected from the groupconsisting of AD-1726057; AD-1725763; AD-1725777; AD-1725057;AD-1725096; AD-1728786; AD-1725059; AD-1728276; AD-1728278; AD-1726936;AD-1725472; AD-1724715; AD-1727292; AD-1730477; AD-1727288; AD-1730167;AD-1725408; and AD-1725761.

In one embodiment, the sense strand and the antisense strand comprise atleast 15, e.g., 15, 16, 17, 18, 19, 20, 21, 22, or 23, contiguousnucleotides differing by no more than 0, 1, 2, 3, or 4 nucleotides fromany one of the sense strand and the antisense strand nucleotidesequences of a duplex selected from the group consisting of AD-1726057;AD-1725763; AD-1725777; AD-1725057; AD-1725096; AD-1728786; AD-1725059;AD-1728276; AD-1728278; AD-1726936; AD-1725472; AD-1724715; AD-1727292;AD-1730477; AD-1727288; AD-1730167; AD-1725408; and AD-1725761.

In one embodiment, the sense strand and the antisense strand comprisethe sense strand and the antisense strand nucleotide sequences of aduplex selected from the group consisting of AD-1726057; AD-1725763;AD-1725777; AD-1725057; AD-1725096; AD-1728786; AD-1725059; AD-1728276;AD-1728278; AD-1726936; AD-1725472; AD-1724715; AD-1727292; AD-1730477;AD-1727288; AD-1730167; AD-1725408; and AD-1725761.

In one embodiment, the sense strand and the antisense strand consist ofthe sense strand and the antisense strand nucleotide sequences of aduplex selected from the group consisting of AD-1726057; AD-1725763;AD-1725777; AD-1725057; AD-1725096; AD-1728786; AD-1725059; AD-1728276;AD-1728278; AD-1726936; AD-1725472; AD-1724715; AD-1727292; AD-1730477;AD-1727288; AD-1730167; AD-1725408; and AD-1725761.

In one embodiment, the dsRNA agent comprises at least one modifiednucleotide.

In one embodiment, substantially all of the nucleotides of the sensestrand; substantially all of the nucleotides of the antisense strandcomprise a modification; or substantially all of the nucleotides of thesense strand and substantially all of the nucleotides of the antisensestrand comprise a modification.

In one embodiment, all of the nucleotides of the sense strand comprise amodification; all of the nucleotides of the antisense strand comprise amodification; or all of the nucleotides of the sense strand and all ofthe nucleotides of the antisense strand comprise a modification.

In one embodiment, at least one of the modified nucleotides is selectedfrom the group consisting of a deoxy-nucleotide, a 3′-terminaldeoxythimidine (dT) nucleotide, a 2′-O-methyl modified nucleotide, a2′-fluoro modified nucleotide, a 2′-deoxy-modified nucleotide, a lockednucleotide, an unlocked nucleotide, a conformationally restrictednucleotide, a constrained ethyl nucleotide, an abasic nucleotide, a2′-amino-modified nucleotide, a 2′-O-allyl-modified nucleotide,2′-C-alkyl-modified nucleotide, a 2′-methoxyethyl modified nucleotide, a2′-O-alkyl-modified nucleotide, a morpholino nucleotide, aphosphoramidate, a non-natural base comprising nucleotide, atetrahydropyran modified nucleotide, a 1,5-anhydrohexitol modifiednucleotide, a cyclohexenyl modified nucleotide, a nucleotide comprisinga phosphorothioate group, a nucleotide comprising a methylphosphonategroup, a nucleotide comprising a 5′-phosphate, a nucleotide comprising a5′-phosphate mimic, a nucleotide comprising a 2′-phosphate group, e.g.,cytidine-2′-phosphate (C2p); guanosine-2′-phosphate (G2p);uridine-2′-phosphate (U2p); adenosine-2′-phosphate (A2p); a thermallydestabilizing nucleotide, a glycol modified nucleotide (GNA), and a2-O—(N-methylacetamide) modified nucleotide; and combinations thereof.

In one embodiment, the modifications on the nucleotides are selectedfrom the group consisting of LNA, HNA, CeNA, 2′-methoxyethyl,2′-O-alkyl, 2′-O-allyl, 2′-C-allyl, 2′-fluoro, 2′-deoxy, 2′-hydroxyl,and glycol; and combinations thereof.

In one embodiment, at least one of the modified nucleotides is selectedfrom the group consisting of a deoxy-nucleotide, a 2′-O-methyl modifiednucleotide, a 2′-fluoro modified nucleotide, a 2′-deoxy-modifiednucleotide, a glycol modified nucleotide (GNA), e.g., Ggn, Cgn, Tgn, orAgn, a nucleotide comprising a 2′-phosphate group, and, avinyl-phosphonate nucleotide; and combinations thereof.

In another embodiment, at least one of the modifications on thenucleotides is a thermally destabilizing nucleotide modification.

In one embodiment, the thermally destabilizing nucleotide modificationis selected from the group consisting of an abasic modification; amismatch with the opposing nucleotide in the duplex; and destabilizingsugar modification, a 2′-deoxy modification, an acyclic nucleotide, anunlocked nucleic acid (UNA), and a glycerol nucleic acid (GNA).

The double stranded region may be 19-30 nucleotide pairs in length;19-25 nucleotide pairs in length; 19-23 nucleotide pairs in length;23-27 nucleotide pairs in length; or 21-23 nucleotide pairs in length.

In one embodiment, each strand is independently no more than 30nucleotides in length.

In one embodiment, the sense strand is 21 nucleotides in length and theantisense strand is 23 nucleotides in length.

The region of complementarity may be at least 17 nucleotides in length;19-23 nucleotides in length; or 19 nucleotides in length.

In one embodiment, at least one strand comprises a 3′ overhang of atleast 1 nucleotide. In another embodiment, at least one strand comprisesa 3′ overhang of at least 2 nucleotides.

In one embodiment, the dsRNA agent further comprises a ligand.

In one embodiment, the ligand is conjugated to the 3′ end of the sensestrand of the dsRNA agent.

In one embodiment, the ligand is an N-acetylgalactosamine (GalNAc)derivative.

In one embodiment, the ligand is one or more GalNAc derivatives attachedthrough a monovalent, bivalent, or trivalent branched linker.

In one embodiment, the ligand is

In one embodiment, the dsRNA agent is conjugated to the ligand as shownin the following schematic

and, wherein X is O or S.

In one embodiment, the X is O.

In one embodiment, the dsRNA agent further comprises at least onephosphorothioate or methylphosphonate internucleotide linkage.

In one embodiment, the phosphorothioate or methylphosphonateinternucleotide linkage is at the 3′-terminus of one strand, e.g., theantisense strand or the sense strand.

In another embodiment, the phosphorothioate or methylphosphonateinternucleotide linkage is at the 5′-terminus of one strand, e.g., theantisense strand or the sense strand.

In one embodiment, the phosphorothioate or methylphosphonateinternucleotide linkage is at the both the 5′- and 3′-terminus of onestrand. In one embodiment, the strand is the antisense strand.

In one embodiment, the base pair at the 1 position of the 5′-end of theantisense strand of the duplex is an AU base pair.

The present invention also provides cells containing any of the dsRNAagents of the invention and pharmaceutical compositions comprising anyof the dsRNA agents of the invention.

The pharmaceutical composition of the invention may include dsRNA agentin an unbuffered solution, e.g., saline or water, or the pharmaceuticalcomposition of the invention may include the dsRNA agent is in a buffersolution, e.g., a buffer solution comprising acetate, citrate,prolamine, carbonate, or phosphate or any combination thereof; orphosphate buffered saline (PBS).

In one aspect, the present invention provides a method of inhibitingexpression of a complement factor B (CFB) gene in a cell. The methodincludes contacting the cell with any of the dsRNAs of the invention orany of the pharmaceutical compositions of the invention, therebyinhibiting expression of the CFB gene in the cell.

In one embodiment, the cell is within a subject, e.g., a human subject,e.g., a subject having a complement factor B-associated disorder. Suchdisorders are typically associated with inflammation or immune systemactivation, e.g., membrane attack complex-mediated lysis, anaphylaxis,or hemolysis. Non-limiting examples of complement factor B-associateddisorders include paroxysmal nocturnal hemoglobinuria (PNH), atypicalhemolytic uremic syndrome (aHUS), asthma, rheumatoid arthritis (RA);antiphospholipid antibody syndrome; lupus nephritis;ischemia-reperfusion injury; typical or infectious hemolytic uremicsyndrome (tHUS); dense deposit disease (DDD); neuromyelitis optica(NMO); multifocal motor neuropathy (MMN); multiple sclerosis (MS);macular degeneration (e.g., age-related macular degeneration (AMD));hemolysis, elevated liver enzymes, and low platelets (HELLP) syndrome;thrombotic thrombocytopenic purpura (TTP); spontaneous fetal loss;Pauci-immune vasculitis; epidermolysis bullosa; recurrent fetal loss;pre-eclampsia, traumatic brain injury, myasthenia gravis, coldagglutinin disease, dermatomyositis bullous pemphigoid, Shiga toxin E.coli-related hemolytic uremic syndrome, C3 neuropathy, anti-neutrophilcytoplasmic antibody-associated vasculitis (e.g., granulomatosis withpolyangiitis (previously known as Wegener granulomatosis), Churg-Strausssyndrome, and microscopic polyangiitis), humoral and vascular transplantrejection, graft dysfunction, myocardial infarction (e.g., tissue damageand ischemia in myocardial infarction), an allogenic transplant, sepsis(e.g., poor outcome in sepsis), Coronary artery disease,dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease,systemic inflammatory response sepsis, septic shock, spinal cord injury,glomerulonephritis, Hashimoto's thyroiditis, type I diabetes, psoriasis,pemphigus, autoimmune hemolytic anemia (AIHA), ITP, Goodpasturesyndrome, Degos disease, antiphospholipid syndrome (APS), catastrophicAPS (CAPS), a cardiovascular disorder, myocarditis, a cerebrovasculardisorder, a peripheral (e.g., musculoskeletal) vascular disorder, arenovascular disorder, a mesenteric/enteric vascular disorder,vasculitis, Henoch-Schönlein purpura nephritis, systemic lupuserythematosus-associated vasculitis, vasculitis associated withrheumatoid arthritis, immune complex vasculitis, Takayasu's disease,dilated cardiomyopathy, diabetic angiopathy, Kawasaki's disease(arteritis), venous gas embolus (VGE), and restenosis following stentplacement, rotational atherectomy, and percutaneous transluminalcoronary angioplasty (PTCA) (see, e.g., Holers (2008) ImmunologicalReviews 223:300-316; Holers and Thurman (2004) Molecular Immunology41:147-152; U.S. Patent Publication No. 20070172483).

In one embodiment, the complement factor B-associate disease is selectedfrom the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, polycystic kidney disease, membranous nephropathy,age-related macular degeneration, atypical hemolytic uremic syndrome,thrombotic microangiopathy, myasthenia gravis, ischemia and reperfusioninjury, paroxysmal nocturnal hemoglobinuria, and rheumatoid arthritis

In another embodiment, the complement factor B-associate disease isselected from the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

In one embodiment, contacting the cell with the dsRNA agent inhibits theexpression of CFB by at least 50%, 60%, 70%, 80%, 90%, or 95%.

In one embodiment, inhibiting expression of CFB decreases CFB proteinlevel in serum of the subject by at least 50%, 60%, 70%, 80%, 90%, or95%.

In one aspect, the present invention provides a method of treating asubject having a disorder that would benefit from reduction incomplement factor B (CFB) expression. The method includes administeringto the subject a therapeutically effective amount of any of the dsRNAsof the invention or any of the pharmaceutical compositions of theinvention, thereby treating the subject having the disorder that wouldbenefit from reduction in CFB expression.

In another aspect, the present invention provides a method of preventingdevelopment of a disorder that would benefit from reduction incomplement factor B (CFB) expression in a subject having at least onesign or symptom of a disorder who does not yet meet the diagnosticcriteria for that disorder. The method includes administering to thesubject a prophylactically effective amount of any of the dsRNAs of theinvention or any of the pharmaceutical compositions of the invention,thereby preventing the subject progressing to meet the diagnosticcriteria of the disorder that would benefit from reduction in CFBexpression.

In one embodiment, the disorder is a complement factorB-(CFB)-associated disorder.

In one embodiment, the subject is human.

In one embodiment, the dsRNA agent is administered to the subject at adose of about 0.01 mg/kg to about 50 mg/kg.

In one embodiment, the dsRNA agent is administered to the subjectsubcutaneously.

In one embodiment, the level of CFB in the subject sample(s) is a CFBprotein level in a blood or serum sample(s).

In one embodiment, the administration of the agent to the subject causesa decrease in hemolysis or a decrease in CFB protein accumulation.

In certain embodiments, the methods of the invention further compriseadministering to the subject an additional therapeutic agent.

In some aspects, the additional therapeutic agent is an iRNA agenttargeting a C5 gene, such as those described in U.S. Pat. No. 9,249,415,the entire contents of which are hereby incorporated herein byreference.

In other aspects, the additional therapeutic agent is an iRNA agenttargeting a complement factor B (CFB) gene, such as those described inU.S. Pat. No. 10,465,194, the entire contents of which are herebyincorporated herein by reference.

In other aspects, the additional therapeutic agent is an inhibitor ofC5, such as an anti-complement component C5 antibody, or antigen-bindingfragment thereof (e.g., eculizumab, ravulizumab-cwvz, or pozelimab(REGN3918)) or a C5 peptide inhibitor (e.g., zilucoplan). Eculizumab isa humanized monoclonal IgG2/4, kappa light chain antibody thatspecifically binds complement component C5 with high affinity andinhibits cleavage of C5 to C5a and C5b, thereby inhibiting thegeneration of the terminal complement complex C5b-9. Eculizumab isdescribed in U.S. Pat. No. 6,355,245, the entire contents of which areincorporated herein by reference. Ravulizumab-cwvz is a humanized IgG2/4monoclonal antibody that specifically binds complement component C5 withhigh affinity and inhibits cleavage of C5 to C5a and C5b, therebyinhibiting the generation of the terminal complement complex C5b-9.Ravulizumab-cwvz is described in WO2015134894, the entire contents ofwhich are incorporated herein by reference. Pozelimab (also known asH4H12166P, described in US20170355757, the entire contents of which areincorporated herein by reference) is a fully-human IgG4 monoclonalantibody designed to block complement factor C5. Zilucoplan is asynthetic, macrocyclic peptide that binds complement component 5 (C5)with sub-nanomolar affinity and allosterically inhibits its cleavageinto C5a and C5b upon activation of the classical, alternative, orlectin pathways (see, e.g., WO2017105939, the entire contexts of whichare incorporated herein by reference).

In yet other aspects, the additional therapeutic is a C3 peptideinhibitor, or analog thereof. In one embodiment, the C3 peptideinhibitor is compstatin. Compstatin is a cyclic tridecapeptide withpotent and selective C3 inhibitory activity. Compstatin, and itsanalogs, are described in U.S. Pat. Nos. 7,888,323, 7,989,589, and8,442,776, in U.S. Patent Publication No. 2012/0178694 and 2013/0053302,and in PCT Publication Nos. WO 2012/174055, WO 2012/2178083, WO2013/036778, the entire contents of each of which are incorporatedherein by reference.

In certain embodiments, treatments known in the art for the variousCFB-associated diseases are used in combination with the RNAi agents ofthe invention.

The present invention also provides kits comprising any of the dsRNAs ofthe invention or any of the pharmaceutical compositions of theinvention, and optionally, instructions for use.

The present invention further provides an RNA-induced silencing complex(RISC) comprising an antisense strand of any of the dsRNA agents of theinvention.

In another embodiment, the RNAi agent is a pharmaceutically acceptablesalt thereof. “Pharmaceutically acceptable salts” of each of RNAi agentsherein include, but are not limited to, a sodium salt, a calcium salt, alithium salt, a potassium salt, an ammonium salt, a magnesium salt, anmixtures thereof. One skilled in the art will appreciate that the RNAiagent, when provided as a polycationic salt having one cation per freeacid group of the optionally modified phosphodiester backbone and/or anyother acidic modifications (e.g., 5′-terminal phosphonate groups). Forexample, an oligonucleotide of “n” nucleotides in length contains n-1optionally modified phosphodiesters, so that an oligonucleotide of 21 ntin length may be provided as a salt having up to 20 cations (e.g, 20sodium cations). Similarly, an RNAi agents having a sense strand of 21nt in length and an antisense strand of 23 nt in length may be providedas a salt having up to 42 cations (e.g, 42 sodium cations). In thepreceding example, where the RNAi agent also includes a 5′-terminalphosphate or a 5′-terminal vinylphosphonate group, the RNAi agent may beprovided as a salt having up to 44 cations (e.g, 44 sodium cations).

The present invention is further illustrated by the following detaileddescription and drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts the three complement pathways: alternative, classical andlectin.

FIG. 2 is a graph depicting the level of human CFB protein in mice (n=3per group) subcutaneously administered a single 1 mg/kg dose of theindicated dsRNA duplexes on day 7. Human CFB protein levels were shownrelative to control levels determined with PBS treatment.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides iRNA compositions which effect theRNA-induced silencing complex (RISC)-mediated cleavage of RNAtranscripts of a complement factor B (CFB) gene. The gene may be withina cell, e.g., a cell within a subject, such as a human. The use of theseiRNAs enables the targeted degradation of mRNAs of the correspondinggene (complement factor B gene) in mammals.

The iRNAs of the invention have been designed to target the humancomplement factor B gene, including portions of the gene that areconserved in the complement factor B orthologs of other mammalianspecies. Without intending to be limited by theory, it is believed thata combination or sub-combination of the foregoing properties and thespecific target sites or the specific modifications in these iRNAsconfer to the iRNAs of the invention improved efficacy, stability,potency, durability, and safety.

Accordingly, the present invention provides methods for treating andpreventing a complement factor B-associated disorder, disease, orcondition, e.g., a disorder, disease, or condition with inflammation orimmune system activation, e.g., membrane attack complex-mediated lysis,anaphylaxis, or hemolysis, e.g., C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease, using iRNA compositionswhich effect the RNA-induced silencing complex (RISC)-mediated cleavageof RNA transcripts of a complement factor B gene.

The iRNAs of the invention include an RNA strand (the antisense strand)having a region which is up to about 30 nucleotides or less in length,e.g., 19-30, 19-29, 19-28, 19-27, 19-26, 19-25, 19-24, 19-23, 19-22,19-21, 19-20, 20-30, 20-29, 20-28, 20-27, 20-26, 20-25, 20-24,20-23,20-22, 20-21, 21-30, 21-29, 21-28, 21-27, 21-26, 21-25, 21-24, 21-23, or21-22 nucleotides in length, which region is substantially complementaryto at least part of an mRNA transcript of a complement factor B gene. Incertain embodiments, the RNAi agents of the disclosure include an RNAstrand (the antisense strand) having a region which is about 21-23nucleotides in length, which region is substantially complementary to atleast part of an mRNA transcript of a complement factor B gene.

In certain embodiments, one or both of the strands of the doublestranded RNAi agents of the invention is up to 66 nucleotides in length,e.g., 36-66, 26-36, 25-36, 31-60, 22-43, 27-53 nucleotides in length,with a region of at least 19 contiguous nucleotides that issubstantially complementary to at least a part of an mRNA transcript ofa complement factor B gene. In some embodiments, such iRNA agents havinglonger length antisense strands may include a second RNA strand (thesense strand) of 20-60 nucleotides in length wherein the sense andantisense strands form a duplex of 18-30 contiguous nucleotides.

The use of iRNAs of the invention enables the targeted degradation ofmRNAs of the corresponding gene (complement factor B gene) in mammals.Using in vitro and in vivo assays, the present inventors havedemonstrated that iRNAs targeting a complement factor B gene canpotently mediate RNAi, resulting in significant inhibition of expressionof a complement factor B gene. Thus, methods and compositions includingthese iRNAs are useful for treating a subject having a complement factorB-associated disorder, e.g., C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

Accordingly, the present invention provides methods and combinationtherapies for treating a subject having a disorder that would benefitfrom inhibiting or reducing the expression of a complement factor Bgene, e.g., a complement factor B-associated disease, such as C3glomerulopathy, systemic lupus erythematosus (SLE), e.g., LupusNephritis, IgA nephropathy, diabetic nephropathy, and polycystic kidneydisease, using iRNA compositions which effect the RNA-induced silencingcomplex (RISC)-mediated cleavage of RNA transcripts of a CFB gene.

The present invention also provides methods for preventing at least onesymptom in a subject having a disorder that would benefit frominhibiting or reducing the expression of a complement factor B gene,e.g., C3 glomerulopathy, systemic lupus erythematosus (SLE), e.g., LupusNephritis, IgA nephropathy, diabetic nephropathy, and polycystic kidneydisease.

In certain embodiments, the administration of the dsRNA to the subjectcauses a decrease in CFB mRNA level, CFB protein level, CH₅₀ activity (ameasure of total hemolytic complement), AH₅₀ (a measure the hemolyticactivity of the alternate pathway of complement), lactate dehydrogenase(LDH) (a measure of intravascular hemolysis), hemoglobin levels; thelevel of any one or more of C3, C9, C5, C5a, C5b, and soluble C5b-9complex.

The following detailed description discloses how to make and usecompositions containing iRNAs to inhibit the expression of a complementfactor B gene as well as compositions, uses, and methods for treatingsubjects that would benefit from inhibition or reduction of theexpression of a complement factor B gene, e.g., subjects susceptible toor diagnosed with a complement factor B-associated disorder.

I. Definitions

In order that the present invention may be more readily understood,certain terms are first defined. In addition, it should be noted thatwhenever a value or range of values of a parameter are recited, it isintended that values and ranges intermediate to the recited values arealso intended to be part of this invention.

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element, e.g., a plurality of elements.

The term “including” is used herein to mean, and is used interchangeablywith, the phrase “including but not limited to”.

The term “or” is used herein to mean, and is used interchangeably with,the term “and/or,” unless context clearly indicates otherwise. Forexample, “sense strand or antisense strand” is understood as “sensestrand or antisense strand or sense strand and antisense strand.”

The term “about” is used herein to mean within the typical ranges oftolerances in the art. For example, “about” can be understood as about 2standard deviations from the mean. In certain embodiments, aboutmeans±10%. In certain embodiments, about means±5%. When about is presentbefore a series of numbers or a range, it is understood that “about” canmodify each of the numbers in the series or range.

The term “at least”, “no less than” or “or more” prior to a number orseries of numbers is understood to include the number adjacent to theterm “at least”, and all subsequent numbers or integers that couldlogically be included, as clear from context. For example, the number ofnucleotides in a nucleic acid molecule must be an integer. For example,“at least 19 nucleotides of a 21 nucleotide nucleic acid molecule” meansthat 19, 20, or 21 nucleotides have the indicated property. When atleast is present before a series of numbers or a range, it is understoodthat “at least” can modify each of the numbers in the series or range.

As used herein, “no more than” or “or less” is understood as the valueadjacent to the phrase and logical lower values or integers, as logicalfrom context, to zero. For example, a duplex with an overhang of “nomore than 2 nucleotides” has a 2, 1, or 0 nucleotide overhang. When “nomore than” is present before a series of numbers or a range, it isunderstood that “no more than” can modify each of the numbers in theseries or range. As used herein, ranges include both the upper and lowerlimit.

As used herein, methods of detection can include determination that theamount of analyte present is below the level of detection of the method.

In the event of a conflict between an indicated target site and thenucleotide sequence for a sense or antisense strand, the indicatedsequence takes precedence.

In the event of a conflict between a sequence and its indicated site ona transcript or other sequence, the nucleotide sequence recited in thespecification takes precedence.

As used herein, the term “Complement Factor B,” used interchangeablywith the term “CFB,” refers to the well-known gene and polypeptide, alsoknown in the art as AHUS, BF, CFAB, BFD, FB, GBG, FBI12, B-Factor,Properdin, H2-Bf, Glycine-Rich Beta Glycoprotein, C3 Proaccelerator,Properdin Factor 2B, C3 Proactivator, PBF2, Glycine-RichBeta-Glycoprotein, C3/C5 Convertase, EC 3.4.21, and EC 3.4.21.473.

The term “CFB” includes human CFB, the amino acid and nucleotidesequence of which may be found in, for example, GenBank Accession No.GI:1732746151; mouse CFB, the amino acid and nucleotide sequence ofwhich may be found in, for example, GenBank Accession Nos. GI:218156288and GI:218156290; rat CFB, the amino acid and nucleotide sequence ofwhich may be found in, for example, GenBank Accession No. GI:218156284;and chimpanzee CFB, the amino acid and nucleotide sequence of which maybe found in, for example, GenBank Accession No. GI:57114201. The term“CFB” also includes Macaca fascicularis CFB, the amino acid andnucleotide sequence of which may be found in, for example, GenBankAccession No. GI:544428919 and in the entry for the gene,ENSMMUP00000000985 (locus=scaffold3881:47830:53620), in the Macacagenome project web site(macaque.genomics.org.cn/page/species/index.jsp). Additional examples ofCFB mRNA sequences are readily available using, e.g., GenBank, UniProt,OMIM, and the Macaca genome project web site. Exemplary CFB nucleotidesequences may also be found in SEQ ID NOs:1-7. SEQ ID NOs:8-14 are theantisense sequences of SEQ ID NOs: 1-7, respectively.

The term “CFB,” as used herein, also refers to naturally occurring DNAsequence variations of the CFB gene. Non-limiting examples of sequencevariations within the CFB gene include 1598A>G in exon 12, which resultsin a lysine being changed to an arginine at amino acid residue 533;858C>G in exon 6, which results in a phenylalanine being changed to aleucine at amino acid residue 286; and 967A>G in exon 7, which resultsin a lysine being changed to an alanine at amino acid residue 323(Tawadrous H. et al. (2010) Pediatr Nephrol. 25:947; Goicoechea de JorgeE et al. (2007) Proc Natl Acad Sci. USA 104:240). The term “CFB,” asused herein, also refers to single nucleotide polymorphisms in the CFBgene. Numerous sequence variations within the CFB gene have beenidentified and may be found at, for example, NCBI dbSNP and UniProt(see, e.g., ncbi.nlm.nih.gov/snp).

Further information on CFB can be found, for example, atwww.ncbi.nlm.nih.gov/gene/629.

Additional examples of CFB mRNA sequences are readily available throughpublicly available databases, e.g., GenBank, UniProt, OMIM, and theMacaca genome project web site.

The entire contents of each of the foregoing GenBank Accession numbersand the Gene database numbers are incorporated herein by reference as ofthe date of filing this application.

As used herein, “target sequence” refers to a contiguous portion of thenucleotide sequence of an mRNA molecule formed during the transcriptionof a complement factor B gene, including mRNA that is a product of RNAprocessing of a primary transcription product. The target portion of thesequence will be at least long enough to serve as a substrate foriRNA-directed cleavage at or near that portion of the nucleotidesequence of an mRNA molecule formed during the transcription of a CFBgene. In one embodiment, the target sequence is within the proteincoding region of CFB.

The target sequence may be from about 19-36 nucleotides in length, e.g.,about 19-30 nucleotides in length. For example, the target sequence canbe about 19-30 nucleotides, 19-30, 19-29, 19-28, 19-27, 19-26, 19-25,19-24, 19-23, 19-22, 19-21, 19-20, 20-30, 20-29, 20-28, 20-27, 20-26,20-25, 20-24, 20-23, 20-22, 20-21, 21-30, 21-29, 21-28, 21-27, 21-26,21-25, 21-24, 21-23, or 21-22 nucleotides in length. In someembodiments, the target sequence is about 19 to about 30 nucleotides inlength. In other embodiments, the target sequence is about 19 to about25 nucleotides in length. In still other embodiments, the targetsequence is about 19 to about 23 nucleotides in length. In someembodiments, the target sequence is about 21 to about 23 nucleotides inlength. Ranges and lengths intermediate to the above recited ranges andlengths are also contemplated to be part of the invention.

As used herein, the term “strand comprising a sequence” refers to anoligonucleotide comprising a chain of nucleotides that is described bythe sequence referred to using the standard nucleotide nomenclature.

“G,” “C,” “A,” “T,” and “U” each generally stand for a nucleotide thatcontains guanine, cytosine, adenine, thymidine, and uracil as a base,respectively. However, it will be understood that the term“ribonucleotide” or “nucleotide” can also refer to a modifiednucleotide, as further detailed below, or a surrogate replacement moiety(see, e.g., Table 1). The skilled person is well aware that guanine,cytosine, adenine, and uracil can be replaced by other moieties withoutsubstantially altering the base pairing properties of an oligonucleotidecomprising a nucleotide bearing such replacement moiety. For example,without limitation, a nucleotide comprising inosine as its base can basepair with nucleotides containing adenine, cytosine, or uracil. Hence,nucleotides containing uracil, guanine, or adenine can be replaced inthe nucleotide sequences of dsRNA featured in the invention by anucleotide containing, for example, inosine. In another example, adenineand cytosine anywhere in the oligonucleotide can be replaced withguanine and uracil, respectively to form G-U Wobble base pairing withthe target mRNA. Sequences containing such replacement moieties aresuitable for the compositions and methods featured in the invention.

The terms “iRNA”, “RNAi agent,” “iRNA agent,” “RNA interference agent”as used interchangeably herein, refer to an agent that contains RNA asthat term is defined herein, and which mediates the targeted cleavage ofan RNA transcript via an RNA-induced silencing complex (RISC) pathway.iRNA directs the sequence-specific degradation of mRNA through a processknown as RNA interference (RNAi). The iRNA modulates, e.g., inhibits,the expression of a complement factor B gene in a cell, e.g., a cellwithin a subject, such as a mammalian subject.

In one embodiment, an RNAi agent of the invention includes a singlestranded RNA that interacts with a target RNA sequence, e.g., acomplement factor B target mRNA sequence, to direct the cleavage of thetarget RNA. Without wishing to be bound by theory it is believed thatlong double stranded RNA introduced into cells is broken down into siRNAby a Type III endonuclease known as Dicer (Sharp et al. (2001) GenesDev. 15:485). Dicer, a ribonuclease-III-like enzyme, processes the dsRNAinto 19-23 base pair short interfering RNAs with characteristic two base3′ overhangs (Bernstein, et al., (2001) Nature 409:363). The siRNAs arethen incorporated into an RNA-induced silencing complex (RISC) where oneor more helicases unwind the siRNA duplex, enabling the complementaryantisense strand to guide target recognition (Nykanen, et al., (2001)Cell 107:309). Upon binding to the appropriate target mRNA, one or moreendonucleases within the RISC cleave the target to induce silencing(Elbashir, et al., (2001) Genes Dev. 15:188). Thus, in one aspect theinvention relates to a single stranded RNA (siRNA) generated within acell and which promotes the formation of a RISC complex to effectsilencing of the target gene, i.e., a complement factor B (CFB) gene.Accordingly, the term “siRNA” is also used herein to refer to an iRNA asdescribed above.

In certain embodiments, the RNAi agent may be a single-stranded siRNA(ssRNAi) that is introduced into a cell or organism to inhibit a targetmRNA. Single-stranded RNAi agents bind to the RISC endonuclease,Argonaute 2, which then cleaves the target mRNA. The single-strandedsiRNAs are generally 15-30 nucleotides and are chemically modified. Thedesign and testing of single-stranded siRNAs are described in U.S. Pat.No. 8,101,348 and in Lima et al., (2012) Cell 150:883-894, the entirecontents of each of which are hereby incorporated herein by reference.Any of the antisense nucleotide sequences described herein may be usedas a single-stranded siRNA as described herein or as chemically modifiedby the methods described in Lima et al., (2012) Cell 150:883-894.

In certain embodiments, an “iRNA” for use in the compositions, uses, andmethods of the invention is a double stranded RNA and is referred toherein as a “double stranded RNA agent,” “double stranded RNA (dsRNA)molecule,” “dsRNA agent,” or “dsRNA”. The term “dsRNA”, refers to acomplex of ribonucleic acid molecules, having a duplex structurecomprising two anti-parallel and substantially complementary nucleicacid strands, referred to as having “sense” and “antisense” orientationswith respect to a target RNA, i.e., a complement factor B (CFB) gene. Insome embodiments of the invention, a double stranded RNA (dsRNA)triggers the degradation of a target RNA, e.g., an mRNA, through apost-transcriptional gene-silencing mechanism referred to herein as RNAinterference or RNAi.

As used herein, the term “modified nucleotide” refers to a nucleotidehaving, independently, a modified sugar moiety, a modifiedinternucleotide linkage, or modified nucleobase, or any combinationthereof. Thus, the term modified nucleotide encompasses substitutions,additions or removal of, e.g., a functional group or atom, tointernucleoside linkages, sugar moieties, or nucleobases. Themodifications suitable for use in the agents of the invention includeall types of modifications disclosed herein or known in the art. Anysuch modifications, as used in a siRNA type molecule, are encompassed by“iRNA” or “RNAi agent” for the purposes of this specification andclaims.

In certain embodiments of the instant disclosure, inclusion of adeoxy-nucleotide—which is acknowledged as a naturally occurring form ofnucleotide—if present within a RNAi agent can be considered toconstitute a modified nucleotide.

The duplex region may be of any length that permits specific degradationof a desired target RNA through a RISC pathway, and may range from about19 to 36 base pairs in length, e.g., about 19-30 base pairs in length,for example, about 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36 base pairsin length, such as about 19-30, 19-29, 19-28, 19-27, 19-26, 19-25,19-24, 19-23, 19-22, 19-21, 19-20, 20-30, 20-29, 20-28, 20-27, 20-26,20-25, 20-24,20-23, 20-22, 20-21, 21-30, 21-29, 21-28, 21-27, 21-26,21-25, 21-24, 21-23, or 21-22 base pairs in length. In certainembodiments, the duplex region is 19-21 base pairs in length, e.g., 21base pairs in length. Ranges and lengths intermediate to the aboverecited ranges and lengths are also contemplated to be part of thedisclosure.

The two strands forming the duplex structure may be different portionsof one larger RNA molecule, or they may be separate RNA molecules. Wherethe two strands are part of one larger molecule, and therefore areconnected by an uninterrupted chain of nucleotides between the 3′-end ofone strand and the 5′-end of the respective other strand forming theduplex structure, the connecting RNA chain is referred to as a “hairpinloop.” A hairpin loop can comprise at least one unpaired nucleotide. Insome embodiments, the hairpin loop can comprise at least 4, 5, 6, 7, 8,9, 10, 20, 23 or more unpaired nucleotides. In some embodiments, thehairpin loop can be 10 or fewer nucleotides.

In some embodiments, the hairpin loop can be 8 or fewer unpairednucleotides. In some embodiments, the hairpin loop can be 4-10 unpairednucleotides. In some embodiments, the hairpin loop can be 4-8nucleotides.

In certain embodiment, the two strands of double-stranded oligomericcompound can be linked together. The two strands can be linked to eachother at both ends, or at one end only. By linking at one end is meantthat 5′-end of first strand is linked to the 3′-end of the second strandor 3′-end of first strand is linked to 5′-end of the second strand. Whenthe two strands are linked to each other at both ends, 5′-end of firststrand is linked to 3′-end of second strand and 3′-end of first strandis linked to 5′-end of second strand. The two strands can be linkedtogether by an oligonucleotide linker including, but not limited to,(N)n; wherein N is independently a modified or unmodified nucleotide andn is 3-23. In some embodiments, n is 3-10, e.g., 3, 4, 5, 6, 7, 8, 9, or10. In some embodiments, the oligonucleotide linker is selected from thegroup consisting of GNRA, (G)4, (U)4, and (dT)4, wherein N is a modifiedor unmodified nucleotide and R is a modified or unmodified purinenucleotide. Some of the nucleotides in the linker can be involved inbase-pair interactions with other nucleotides in the linker. The twostrands can also be linked together by a non-nucleosidic linker, e.g. alinker described herein. It will be appreciated by one of skill in theart that any oligonucleotide chemical modifications or variationsdescribe herein can be used in the oligonucleotide linker.

Hairpin and dumbbell type oligomeric compounds will have a duplex regionequal to or at least 14, 15, 15, 16, 17, 18, 19, 29, 21, 22, 23, 24, or25 nucleotide pairs. The duplex region can be equal to or less than 200,100, or 50, in length. In some embodiments, ranges for the duplex regionare 15-30, 17 to 23, 19 to 23, and 19 to 21 nucleotides pairs in length.

The hairpin oligomeric compounds can have a single strand overhang orterminal unpaired region, in some embodiments at the 3′, and in someembodiments on the antisense side of the hairpin. In some embodiments,the overhangs are 1-4, more generally 2-3 nucleotides in length. Thehairpin oligomeric compounds that can induce RNA interference are alsoreferred to as “shRNA” herein.

Where the two substantially complementary strands of a dsRNA arecomprised by separate RNA molecules, those molecules need not be, butcan be covalently connected. Where the two strands are connectedcovalently by means other than an uninterrupted chain of nucleotidesbetween the 3′-end of one strand and the 5′-end of the respective otherstrand forming the duplex structure, the connecting structure isreferred to as a “linker.” The RNA strands may have the same or adifferent number of nucleotides. The maximum number of base pairs is thenumber of nucleotides in the shortest strand of the dsRNA minus anyoverhangs that are present in the duplex. In addition to the duplexstructure, an RNAi may comprise one or more nucleotide overhangs. In oneembodiment of the RNAi agent, at least one strand comprises a 3′overhang of at least 1 nucleotide. In another embodiment, at least onestrand comprises a 3′ overhang of at least 2 nucleotides, e.g., 2, 3, 4,5, 6, 7, 9, 10, 11, 12, 13, 14, or 15 nucleotides. In other embodiments,at least one strand of the RNAi agent comprises a 5′ overhang of atleast 1 nucleotide. In certain embodiments, at least one strandcomprises a 5′ overhang of at least 2 nucleotides, e.g., 2, 3, 4, 5, 6,7, 9, 10, 11, 12, 13, 14, or 15 nucleotides. In still other embodiments,both the 3′ and the 5′ end of one strand of the RNAi agent comprise anoverhang of at least 1 nucleotide.

In certain embodiments, an iRNA agent of the invention is a dsRNA, eachstrand of which comprises 19-23 nucleotides, that interacts with atarget RNA sequence, e.g., a complement factor B (CFB) gene, to directcleavage of the target RNA.

In some embodiments, an iRNA of the invention is a dsRNA of 24-30nucleotides that interacts with a target RNA sequence, e.g., a CFBtarget mRNA sequence, to direct the cleavage of the target RNA.

As used herein, the term “nucleotide overhang” refers to at least oneunpaired nucleotide that protrudes from the duplex structure of a doublestranded iRNA. For example, when a 3′-end of one strand of a dsRNAextends beyond the 5′-end of the other strand, or vice versa, there is anucleotide overhang. A dsRNA can comprise an overhang of at least onenucleotide; alternatively, the overhang can comprise at least twonucleotides, at least three nucleotides, at least four nucleotides, atleast five nucleotides or more. A nucleotide overhang can comprise orconsist of a nucleotide/nucleoside analog, including adeoxynucleotide/nucleoside. The overhang(s) can be on the sense strand,the antisense strand, or any combination thereof. Furthermore, thenucleotide(s) of an overhang can be present on the 5′-end, 3′-end, orboth ends of either an antisense or sense strand of a dsRNA.

In one embodiment of the dsRNA, at least one strand comprises a 3′overhang of at least 1 nucleotide. In another embodiment, at least onestrand comprises a 3′ overhang of at least 2 nucleotides, e.g., 2, 3, 4,5, 6, 7, 9, 10, 11, 12, 13, 14, or 15 nucleotides. In other embodiments,at least one strand of the RNAi agent comprises a 5′ overhang of atleast 1 nucleotide. In certain embodiments, at least one strandcomprises a 5′ overhang of at least 2 nucleotides, e.g., 2, 3, 4, 5, 6,7, 9, 10, 11, 12, 13, 14, or 15 nucleotides. In still other embodiments,both the 3′ and the 5′ end of one strand of the RNAi agent comprise anoverhang of at least 1 nucleotide.

In one embodiment, the antisense strand of a dsRNA has a 1-10nucleotide, e.g., a 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotide,overhang at the 3′-end or the 5′-end. In one embodiment, the sensestrand of a dsRNA has a 1-10 nucleotide, e.g., a 1, 2, 3, 4, 5, 6, 7, 8,9, or 10 nucleotide, overhang at the 3′-end or the 5′-end. In anotherembodiment, one or more of the nucleotides in the overhang is replacedwith a nucleoside thiophosphate.

In certain embodiments, the antisense strand of a dsRNA has a 1-10nucleotides, e.g., a 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotide,overhang at the 3′-end or the 5′-end. In certain embodiments, theoverhang on the sense strand or the antisense strand, or both, caninclude extended lengths longer than 10 nucleotides, e.g., 1-30nucleotides, 2-30 nucleotides, 10-30 nucleotides, 10-25 nucleotides,10-20 nucleotides, or 10-15 nucleotides in length. In certainembodiments, an extended overhang is on the sense strand of the duplex.In certain embodiments, an extended overhang is present on the 3′ end ofthe sense strand of the duplex. In certain embodiments, an extendedoverhang is present on the 5′ end of the sense strand of the duplex. Incertain embodiments, an extended overhang is on the antisense strand ofthe duplex. In certain embodiments, an extended overhang is present onthe 3′end of the antisense strand of the duplex. In certain embodiments,an extended overhang is present on the 5′end of the antisense strand ofthe duplex. In certain embodiments, one or more of the nucleotides inthe extended overhang is replaced with a nucleoside thiophosphate. Incertain embodiments, the overhang includes a self-complementary portionsuch that the overhang is capable of forming a hairpin structure that isstable under physiological conditions.

“Blunt” or “blunt end” means that there are no unpaired nucleotides atthat end of the double stranded RNA agent, i.e., no nucleotide overhang.A “blunt ended” double stranded RNA agent is double stranded over itsentire length, i.e., no nucleotide overhang at either end of themolecule. The RNAi agents of the invention include RNAi agents with nonucleotide overhang at one end (i.e., agents with one overhang and oneblunt end) or with no nucleotide overhangs at either end. Most oftensuch a molecule will be double-stranded over its entire length.

The term “antisense strand” or “guide strand” refers to the strand of aniRNA, e.g., a dsRNA, which includes a region that is substantiallycomplementary to a target sequence, e.g., a CFB mRNA.

As used herein, the term “region of complementarity” refers to theregion on the antisense strand that is substantially complementary to asequence, for example a target sequence, e.g., a complement factor Bnucleotide sequence, as defined herein. Where the region ofcomplementarity is not fully complementary to the target sequence, themismatches can be in the internal or terminal regions of the molecule.Generally, the most tolerated mismatches are in the terminal regions,e.g., within 5, 4, or 3 nucleotides of the 5′- or 3′-end of the iRNA. Insome embodiments, a double stranded RNA agent of the invention includesa nucleotide mismatch in the antisense strand. In some embodiments, theantisense strand of the double stranded RNA agent of the inventionincludes no more than 4 mismatches with the target mRNA, e.g., theantisense strand includes 4, 3, 2, 1, or 0 mismatches with the targetmRNA. In some embodiments, the antisense strand double stranded RNAagent of the invention includes no more than 4 mismatches with the sensestrand, e.g., the antisense strand includes 4, 3, 2, 1, or 0 mismatcheswith the sense strand. In some embodiments, a double stranded RNA agentof the invention includes a nucleotide mismatch in the sense strand. Insome embodiments, the sense strand of the double stranded RNA agent ofthe invention includes no more than 4 mismatches with the antisensestrand, e.g., the sense strand includes 4, 3, 2, 1, or 0 mismatches withthe antisense strand. In some embodiments, the nucleotide mismatch is,for example, within 5, 4, 3 nucleotides from the 3′-end of the iRNA. Inanother embodiment, the nucleotide mismatch is, for example, in the3′-terminal nucleotide of the iRNA agent. In some embodiments, themismatch(s) is not in the seed region.

Thus, an RNAi agent as described herein can contain one or moremismatches to the target sequence. In one embodiment, a RNAi agent asdescribed herein contains no more than 3 mismatches (i.e., 3, 2, 1, or 0mismatches). In one embodiment, an RNAi agent as described hereincontains no more than 2 mismatches. In one embodiment, an RNAi agent asdescribed herein contains no more than 1 mismatch. In one embodiment, anRNAi agent as described herein contains 0 mismatches. In certainembodiments, if the antisense strand of the RNAi agent containsmismatches to the target sequence, the mismatch can optionally berestricted to be within the last 5 nucleotides from either the 5′- or3′-end of the region of complementarity. For example, in suchembodiments, for a 23 nucleotide RNAi agent, the strand which iscomplementary to a region of a CFB gene, generally does not contain anymismatch within the central 13 nucleotides. The methods described hereinor methods known in the art can be used to determine whether an RNAiagent containing a mismatch to a target sequence is effective ininhibiting the expression of a CFB gene. Consideration of the efficacyof RNAi agents with mismatches in inhibiting expression of a CFB gene isimportant, especially if the particular region of complementarity in aCFB gene is known to have polymorphic sequence variation within thepopulation.

The term “sense strand” or “passenger strand” as used herein, refers tothe strand of an iRNA that includes a region that is substantiallycomplementary to a region of the antisense strand as that term isdefined herein.

As used herein, “substantially all of the nucleotides are modified” arelargely but not wholly modified and can include not more than 5, 4, 3,2, or 1 unmodified nucleotides.

As used herein, the term “cleavage region” refers to a region that islocated immediately adjacent to the cleavage site. The cleavage site isthe site on the target at which cleavage occurs. In some embodiments,the cleavage region comprises three bases on either end of, andimmediately adjacent to, the cleavage site. In some embodiments, thecleavage region comprises two bases on either end of, and immediatelyadjacent to, the cleavage site. In some embodiments, the cleavage sitespecifically occurs at the site bound by nucleotides 10 and 11 of theantisense strand, and the cleavage region comprises nucleotides 11, 12and 13.

As used herein, and unless otherwise indicated, the term“complementary,” when used to describe a first nucleotide sequence inrelation to a second nucleotide sequence, refers to the ability of anoligonucleotide or polynucleotide comprising the first nucleotidesequence to hybridize and form a duplex structure under certainconditions with an oligonucleotide or polynucleotide comprising thesecond nucleotide sequence, as will be understood by the skilled person.Such conditions can be, for example, “stringent conditions”, wherestringent conditions can include: 400 mM NaCl, 40 mM PIPES pH 6.4, 1 mMEDTA, 50° C. or 70° C. for 12-16 hours followed by washing (see, e.g.,“Molecular Cloning: A Laboratory Manual, Sambrook, et al. (1989) ColdSpring Harbor Laboratory Press). Other conditions, such asphysiologically relevant conditions as can be encountered inside anorganism, can apply. The skilled person will be able to determine theset of conditions most appropriate for a test of complementarity of twosequences in accordance with the ultimate application of the hybridizednucleotides.

Complementary sequences within an iRNA, e.g., within a dsRNA asdescribed herein, include base-pairing of the oligonucleotide orpolynucleotide comprising a first nucleotide sequence to anoligonucleotide or polynucleotide comprising a second nucleotidesequence over the entire length of one or both nucleotide sequences.Such sequences can be referred to as “fully complementary” with respectto each other herein. However, where a first sequence is referred to as“substantially complementary” with respect to a second sequence herein,the two sequences can be fully complementary, or they can form one ormore, but generally not more than 5, 4, 3, or 2 mismatched base pairsupon hybridization for a duplex up to 30 base pairs, while retaining theability to hybridize under the conditions most relevant to theirultimate application, e.g., inhibition of gene expression, in vitro orin vivo. However, where two oligonucleotides are designed to form, uponhybridization, one or more single stranded overhangs, such overhangsshall not be regarded as mismatches with regard to the determination ofcomplementarity. For example, a dsRNA comprising one oligonucleotide 21nucleotides in length and another oligonucleotide 23 nucleotides inlength, wherein the longer oligonucleotide comprises a sequence of 21nucleotides that is fully complementary to the shorter oligonucleotide,can yet be referred to as “fully complementary” for the purposesdescribed herein.

“Complementary” sequences, as used herein, can also include, or beformed entirely from, non-Watson-Crick base pairs or base pairs formedfrom non-natural and modified nucleotides, in so far as the aboverequirements with respect to their ability to hybridize are fulfilled.Such non-Watson-Crick base pairs include, but are not limited to, G:UWobble or Hoogsteen base pairing.

The terms “complementary,” “fully complementary” and “substantiallycomplementary” herein can be used with respect to the base matchingbetween the sense strand and the antisense strand of a dsRNA, or betweentwo oligonucleotides or polynucleotides, such as the antisense strand ofa double stranded RNA agent and a target sequence, as will be understoodfrom the context of their use.

As used herein, a polynucleotide that is “substantially complementary toat least part of” a messenger RNA (mRNA) refers to a polynucleotide thatis substantially complementary to a contiguous portion of the mRNA ofinterest (e.g., an mRNA encoding a complement factor B gene). Forexample, a polynucleotide is complementary to at least a part of acomplement factor B mRNA if the sequence is substantially complementaryto a non-interrupted portion of an mRNA encoding a complement factor Bgene.

Accordingly, in some embodiments, the antisense polynucleotidesdisclosed herein are fully complementary to the target CFB sequence.

In other embodiments, the antisense polynucleotides disclosed herein aresubstantially complementary to the target CFB sequence and comprise acontiguous nucleotide sequence which is at least 80% complementary overits entire length to the equivalent region of the nucleotide sequence ofany one of SEQ ID NOs: 1-7 or a fragment of any one of SEQ ID NOs: 1-7,such as about 85%, about 90%, about 91%, about 92%, about 93%, about94%, about 95%, about 96%, about 97%, about 98%, or about 99%complementary.

In some embodiments, the antisense polynucleotides disclosed herein aresubstantially complementary to a fragment of a target CFB sequence andcomprise a contiguous nucleotide sequence which is at least 80%complementary over its entire length to a fragment of SEQ ID NO: 1selected from the group of nucleotides 504-526, 640-662, 641-663,995-1017, 997-1019, 1034-1056, 1141-1163, 1145-1167, 1389-1411,1473-1495, 1826-1848, 1828-1850, 1842-1864, 2242-2264, 2391-2413,2393-2415, 2438-2460, or 2453-2475 of SEQ ID NO: 1, such as about 85%,about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about96%, about 97%, about 98%, or about 99% complementary.

In other embodiments, the antisense polynucleotides disclosed herein aresubstantially complementary to the target CFB sequence and comprise acontiguous nucleotide sequence which is at least about 80% complementaryover its entire length to any one of the sense strand nucleotidesequences in any one of any one of Tables 2-3, or a fragment of any oneof the sense strand nucleotide sequences in any one of Tables 2-3, suchas about 85%, about 90%, about 95%, or fully complementary.

In one embodiment, an RNAi agent of the disclosure includes a sensestrand that is substantially complementary to an antisensepolynucleotide which, in turn, is the same as a target CFB sequence, andwherein the sense strand polynucleotide comprises a contiguousnucleotide sequence which is at least about 80% complementary over itsentire length to the equivalent region of the nucleotide sequence of SEQID NOs: 8-14, or a fragment of any one of SEQ ID NOs:8-14, such as about85%, about 90%, about 95%, or fully complementary.

In some embodiments, an iRNA of the invention includes a sense strandthat is substantially complementary to an antisense polynucleotidewhich, in turn, is complementary to a target complement factor Bsequence, and wherein the sense strand polynucleotide comprises acontiguous nucleotide sequence which is at least about 80% complementaryover its entire length to any one of the antisense strand nucleotidesequences in any one of any one of Tables 2-3, or a fragment of any oneof the antisense strand nucleotide sequences in any one of Tables 2-3,such as about 85%, about 90%, about 95%, or fully complementary.

In some embodiments, the sense and antisense strands are selected fromthe group consisting of AD-1726057; AD-1725763; AD-1725777; AD-1725057;AD-1725096; AD-1728786; AD-1725059; AD-1728276; AD-1728278; AD-1726936;AD-1725472; AD-1724715; AD-1727292; AD-1730477; AD-1727288; AD-1730167;AD-1725408; and AD-1725761.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1726057.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725763.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725777.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725057.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725096.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1728786.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725059.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1728276.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1728278.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1726936.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725472.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1724715.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1727292.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1730477.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1727288.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1730167.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725408.

In one embodiment, the sense and antisense strands are selected fromduplex AD-1725761.

In some embodiments, the double-stranded region of a double-strandediRNA agent is equal to or at least, 17, 18, 19, 20, 21, 22, 23, 23, 24,25, 26, 27, 28, 29, 30 or more nucleotide pairs in length.

In some embodiments, the antisense strand of a double-stranded iRNAagent is equal to or at least 17, 18, 19, 20, 21, 22, 23, 23, 24, 25,26, 27, 28, 29, or 30 nucleotides in length.

In some embodiments, the sense strand of a double-stranded iRNA agent isequal to or at least 17, 18, 19, 20, 21, 22, 23, 23, 24, 25, 26, 27, 28,29, or 30 nucleotides in length.

In one embodiment, the sense and antisense strands of thedouble-stranded iRNA agent are each 18 to 30 nucleotides in length.

In one embodiment, the sense and antisense strands of thedouble-stranded iRNA agent are each 19 to 25 nucleotides in length.

In one embodiment, the sense and antisense strands of thedouble-stranded iRNA agent are each 21 to 23 nucleotides in length.

In one embodiment, the sense strand of the iRNA agent is 21-nucleotidesin length, and the antisense strand is 23-nucleotides in length, whereinthe strands form a double-stranded region of 21 consecutive base pairshaving a 2-nucleotide long single stranded overhangs at the 3-end.

In some embodiments, the majority of nucleotides of each strand areribonucleotides, but as described in detail herein, each or both strandscan also include one or more non-ribonucleotides, e.g., adeoxyribonucleotide or a modified nucleotide. In addition, an “iRNA” mayinclude ribonucleotides with chemical modifications. Such modificationsmay include all types of modifications disclosed herein or known in theart. Any such modifications, as used in an iRNA molecule, areencompassed by “iRNA” for the purposes of this specification and claims.

In certain embodiments of the instant disclosure, inclusion of adeoxy-nucleotide if present within an RNAi agent can be considered toconstitute a modified nucleotide.

In one embodiment, at least partial suppression of the expression of aCFB gene, is assessed by a reduction of the amount of CFB mRNA which canbe isolated from or detected in a first cell or group of cells in whicha CFB gene is transcribed and which has or have been treated such thatthe expression of a CFB gene is inhibited, as compared to a second cellor group of cells substantially identical to the first cell or group ofcells but which has or have not been so treated (control cells). Thedegree of inhibition may be expressed in terms of:

${\frac{\left( {{mRNA}{in}{control}{cells}} \right) - \left( {{mRNA}{in}{treated}{cells}} \right)}{\left( {{mRNA}{in}{control}{cells}} \right)} \cdot 100}\%$

The phrase “contacting a cell with an iRNA,” such as a dsRNA, as usedherein, includes contacting a cell by any possible means. Contacting acell with an iRNA includes contacting a cell in vitro with the iRNA orcontacting a cell in vivo with the iRNA. The contacting may be donedirectly or indirectly. Thus, for example, the iRNA may be put intophysical contact with the cell by the individual performing the method,or alternatively, the iRNA may be put into a situation that will permitor cause it to subsequently come into contact with the cell.

Contacting a cell in vitro may be done, for example, by incubating thecell with the iRNA. Contacting a cell in vivo may be done, for example,by injecting the iRNA into or near the tissue where the cell is located,or by injecting the iRNA into another area, e.g., the bloodstream or thesubcutaneous space, such that the agent will subsequently reach thetissue where the cell to be contacted is located. For example, the iRNAmay contain or be coupled to a ligand, e.g., GalNAc, that directs theiRNA to a site of interest, e.g., the liver. Combinations of in vitroand in vivo methods of contacting are also possible. For example, a cellmay also be contacted in vitro with an iRNA and subsequentlytransplanted into a subject.

In certain embodiments, contacting a cell with an iRNA includes“introducing” or “delivering the iRNA into the cell” by facilitating oreffecting uptake or absorption into the cell. Absorption or uptake of aniRNA can occur through unaided diffusion or active cellular processes,or by auxiliary agents or devices. Introducing an iRNA into a cell maybe in vitro or in vivo. For example, for in vivo introduction, iRNA canbe injected into a tissue site or administered systemically. In vitrointroduction into a cell includes methods known in the art such aselectroporation and lipofection. Further approaches are described hereinbelow or are known in the art.

The term “lipid nanoparticle” or “LNP” is a vesicle comprising a lipidlayer encapsulating a pharmaceutically active molecule, such as anucleic acid molecule, e.g., an iRNA or a plasmid from which an iRNA istranscribed. LNPs are described in, for example, U.S. Pat. Nos.6,858,225, 6,815,432, 8,158,601, and 8,058,069, the entire contents ofwhich are hereby incorporated herein by reference.

As used herein, a “subject” is an animal, such as a mammal, including aprimate (such as a human, a non-human primate, e.g., a monkey, and achimpanzee), a non-primate (such as a rabbit, a sheep, a hamster, aguinea pig, a dog, a rat, or a mouse), or a bird that expresses thetarget gene, either endogenously or heterologously. In an embodiment,the subject is a human, such as a human being treated or assessed for adisease or disorder that would benefit from reduction in CFB expression;a human at risk for a disease or disorder that would benefit fromreduction in CFB expression; a human having a disease or disorder thatwould benefit from reduction in CFB expression; or human being treatedfor a disease or disorder that would benefit from reduction in CFBexpression as described herein. In some embodiments, the subject is afemale human. In other embodiments, the subject is a male human. In oneembodiment, the subject is an adult subject. In another embodiment, thesubject is a pediatric subject.

As used herein, the terms “treating” or “treatment” refer to abeneficial or desired result, such as reducing at least one sign orsymptom of a CFB-associated disorder in a subject. Treatment alsoincludes a reduction of one or more sign or symptoms associated withunwanted CFB expression; diminishing the extent of unwanted CFBactivation or stabilization; amelioration or palliation of unwanted CFBactivation or stabilization. “Treatment” can also mean prolongingsurvival as compared to expected survival in the absence of treatment.

The term “lower” in the context of the level of CFB in a subject or adisease marker or symptom refers to a statistically significant decreasein such level. The decrease can be, for example, at least 10%, 15%, 20%,25%, 30%, %, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%,or more. In certain embodiments, a decrease is at least 20%. In certainembodiments, the decrease is at least 50% in a disease marker, e.g.,protein or gene expression level. “Lower” in the context of the level ofCFB in a subject is a decrease to a level accepted as within the rangeof normal for an individual without such disorder. In certainembodiments, the expression of the target is normalized, i.e., decreasedtowards or to a level accepted as within the range of normal for anindividual without such disorder, e.g., normalization of body weight,blood pressure, or a serum lipid level. As used here, “lower” in asubject can refer to lowering of gene expression or protein productionin a cell in a subject does not require lowering of expression in allcells or tissues of a subject. For example, as used herein, lowering ina subject can include lowering of gene expression or protein productionin the liver of a subject.

The term “lower” can also be used in association with normalizing asymptom of a disease or condition, i.e. decreasing the differencebetween a level in a subject suffering from a CFB-associated diseasetowards or to a level in a normal subject not suffering from aCFB-associated disease. For example, if a subject with a normal weightof 70 kg weighs 90 kg prior to treatment (20 kg overweight) and 80 kgafter treatment (10 kg overweight), the subject's weight is loweredtowards a normal weight by 50% (10/20×100%). Similarly, if the HDL levelof a woman is increased from 50 mg/dL (poor) to 57 mg/dL, with a normallevel being 60 mg/dL, the difference between the prior level of thesubject and the normal level is decreased by 70% (difference of 10 mg/dLbetween subject level and normal is decreased by 7 mg/dL, 7/10×100%). Asused herein, if a disease is associated with an elevated value for asymptom, “normal” is considered to be the upper limit of normal. If adisease is associated with a decreased value for a symptom, “normal” isconsidered to be the lower limit of normal.

As used herein, “prevention” or “preventing,” when used in reference toa disease, disorder or condition thereof, that would benefit from areduction in expression of a CFB gene or production of CFB protein,refers to preventing a subject who has at least one sign or symptom of adisease from developing further signs and symptoms thereby meeting thediagnostic criteria for that disease. In certain embodiments, preventionincludes delayed progression to meeting the diagnostic criteria of thedisease by days, weeks, months or years as compared to what would bepredicted by natural history studies or the typical progression of thedisease.

As used herein, the term “complement factor B disease” or“CFB-associated disease,” is a disease or disorder that is caused by, orassociated with, complement activation. The term “CFB-associateddisease” includes a disease, disorder or condition that would benefitfrom a decrease in CFB gene expression, replication, or proteinactivity. Non-limiting examples of CFB-associated diseases include, forexample, paroxysmal nocturnal hemoglobinuria (PNH), atypical hemolyticuremic syndrome (aHUS), asthma, rheumatoid arthritis (RA);antiphospholipid antibody syndrome; lupus nephritis;ischemia-reperfusion injury; typical or infectious hemolytic uremicsyndrome (tHUS); dense deposit disease (DDD); neuromyelitis optica(NMO); multifocal motor neuropathy (MMN); multiple sclerosis (MS);macular degeneration (e.g., age-related macular degeneration (AMD));hemolysis, elevated liver enzymes, and low platelets (HELLP) syndrome;thrombotic thrombocytopenic purpura (TTP); spontaneous fetal loss;Pauci-immune vasculitis; epidermolysis bullosa; recurrent fetal loss;pre-eclampsia, traumatic brain injury, myasthenia gravis, coldagglutinin disease, dermatomyositis bullous pemphigoid, Shiga toxin E.coli-related hemolytic uremic syndrome, C3 neuropathy, anti-neutrophilcytoplasmic antibody-associated vasculitis (e.g., granulomatosis withpolyangiitis (previously known as Wegener granulomatosis), Churg-Strausssyndrome, and microscopic polyangiitis), humoral and vascular transplantrejection, graft dysfunction, myocardial infarction (e.g., tissue damageand ischemia in myocardial infarction), an allogenic transplant, sepsis(e.g., poor outcome in sepsis), Coronary artery disease,dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease,systemic inflammatory response sepsis, septic shock, spinal cord injury,glomerulonephritis, Hashimoto's thyroiditis, type I diabetes, psoriasis,pemphigus, autoimmune hemolytic anemia (AIHA), ITP, Goodpasturesyndrome, Degos disease, antiphospholipid syndrome (APS), catastrophicAPS (CAPS), a cardiovascular disorder, myocarditis, a cerebrovasculardisorder, a peripheral (e.g., musculoskeletal) vascular disorder, arenovascular disorder, a mesenteric/enteric vascular disorder,vasculitis, Henoch-Schönlein purpura nephritis, systemic lupuserythematosus-associated vasculitis, vasculitis associated withrheumatoid arthritis, immune complex vasculitis, Takayasu's disease,dilated cardiomyopathy, diabetic angiopathy, Kawasaki's disease(arteritis), venous gas embolus (VGE), and restenosis following stentplacement, rotational atherectomy, and percutaneous transluminalcoronary angioplasty (PTCA) (see, e.g., Holers (2008) ImmunologicalReviews 223:300-316; Holers and Thurman (2004) Molecular Immunology41:147-152; U.S. Patent Publication No. 20070172483).

In one embodiment, the complement factor B-associate disease is selectedfrom the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, polycystic kidney disease, membranous nephropathy,age-related macular degeneration, atypical hemolytic uremic syndrome,thrombotic microangiopathy, myasthenia gravis, ischemia and reperfusioninjury, paroxysmal nocturnal hemoglobinuria, and rheumatoid arthritis

In another embodiment, the complement factor B-associate disease isselected from the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

Further details regarding signs and symptoms of the various diseases orconditions are provided herein and are well known in the art.

“Therapeutically effective amount,” as used herein, is intended toinclude the amount of an RNAi agent that, when administered to a subjecthaving a CFB-associated disease, is sufficient to effect treatment ofthe disease (e.g., by diminishing, ameliorating, or maintaining theexisting disease or one or more symptoms of disease). The“therapeutically effective amount” may vary depending on the RNAi agent,how the agent is administered, the disease and its severity and thehistory, age, weight, family history, genetic makeup, the types ofpreceding or concomitant treatments, if any, and other individualcharacteristics of the subject to be treated.

“Prophylactically effective amount,” as used herein, is intended toinclude the amount of an RNAi agent that, when administered to a subjecthaving at least one sign or symptom of a CFB-associated disorder, issufficient to prevent or delay the subject's progression to meeting thefull diagnostic criteria of the disease. Prevention of the diseaseincludes slowing the course of progression to full blown disease. The“prophylactically effective amount” may vary depending on the RNAiagent, how the agent is administered, the degree of risk of disease, andthe history, age, weight, family history, genetic makeup, the types ofpreceding or concomitant treatments, if any, and other individualcharacteristics of the patient to be treated.

A “therapeutically-effective amount” or “prophylactically effectiveamount” also includes an amount of an RNAi agent that produces somedesired effect at a reasonable benefit/risk ratio applicable to anytreatment. The iRNA employed in the methods of the present invention maybe administered in a sufficient amount to produce a reasonablebenefit/risk ratio applicable to such treatment.

The phrase “pharmaceutically acceptable” is employed herein to refer tothose compounds, materials, compositions, or dosage forms which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of human subjects and animal subjects without excessivetoxicity, irritation, allergic response, or other problem orcomplication, commensurate with a reasonable benefit/risk ratio.

The phrase “pharmaceutically-acceptable carrier” as used herein means apharmaceutically-acceptable material (including salts), composition, orvehicle, such as a liquid or solid filler, diluent, excipient,manufacturing aid (e.g., lubricant, talc magnesium, calcium or zincstearate, or steric acid), or solvent encapsulating material, involvedin carrying or transporting the subject compound from one organ, orportion of the body, to another organ, or portion of the body. Eachcarrier must be “acceptable” in the sense of being compatible with theother ingredients of the formulation and not injurious to the subjectbeing treated. Such carriers are known in the art. Pharmaceuticallyacceptable carriers include carriers for administration by injection.

The term “sample,” as used herein, includes a collection of similarfluids, cells, or tissues isolated from a subject, as well as fluids,cells, or tissues present within a subject. Examples of biologicalfluids include blood, serum and serosal fluids, plasma, cerebrospinalfluid, ocular fluids, lymph, urine, saliva, and the like. Tissue samplesmay include samples from tissues, organs, or localized regions. Forexample, samples may be derived from particular organs, parts of organs,or fluids or cells within those organs. In certain embodiments, samplesmay be derived from the liver (e.g., whole liver or certain segments ofliver or certain types of cells in the liver, such as, e.g.,hepatocytes). In some embodiments, a “sample derived from a subject”refers to urine obtained from the subject. A “sample derived from asubject” can refer to blood or blood derived serum or plasma from thesubject.

II. iRNAs of the Invention

The present invention provides iRNAs which inhibit the expression of acomplement factor B gene. In certain embodiments, the iRNA includesdouble stranded ribonucleic acid (dsRNA) molecules for inhibiting theexpression of a CFB gene in a cell, such as a cell within a subject,e.g., a mammal, such as a human susceptible to developing a complementfactor B-associated disorder. The dsRNAi agent includes an antisensestrand having a region of complementarity which is complementary to atleast a part of an mRNA formed in the expression of a CFB gene. Theregion of complementarity is about 19-30 nucleotides in length (e.g.,about 30, 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, or 19 nucleotides inlength). Upon contact with a cell expressing the CFB gene, the iRNAinhibits the expression of the CFB gene (e.g., a human, a primate, anon-primate, or a rat CFB gene) by at least about 50% as assayed by, forexample, a PCR or branched DNA (bDNA)-based method, or by aprotein-based method, such as by immunofluorescence analysis, using, forexample, western blotting or flow cytometric techniques. In certainembodiments, inhibition of expression is determined by the qPCR methodprovided in the examples herein with the siRNA at, e.g., a 10 nMconcentration, in an appropriate organism cell line provided therein. Incertain embodiments, inhibition of expression in vivo is determined byknockdown of the human gene in a rodent expressing the human gene, e.g.,a mouse or an AAV-infected mouse expressing the human target gene, e.g.,when administered as single dose, e.g., at 3 mg/kg at the nadir of RNAexpression.

A dsRNA includes two RNA strands that are complementary and hybridize toform a duplex structure under conditions in which the dsRNA will beused. One strand of a dsRNA (the antisense strand) includes a region ofcomplementarity that is substantially complementary, and generally fullycomplementary, to a target sequence. The target sequence can be derivedfrom the sequence of an mRNA formed during the expression of a CFB gene.The other strand (the sense strand) includes a region that iscomplementary to the antisense strand, such that the two strandshybridize and form a duplex structure when combined under suitableconditions. As described elsewhere herein and as known in the art, thecomplementary sequences of a dsRNA can also be contained asself-complementary regions of a single nucleic acid molecule, as opposedto being on separate oligonucleotides.

Generally, the duplex structure is 15 to 30 base pairs in length, e.g.,15-29, 15-28, 15-27, 15-26, 15-25, 15-24, 15-23, 15-22, 15-21, 15-20,15-19, 15-18, 15-17, 18-30, 18-29, 18-28, 18-27, 18-26, 18-25, 18-24,18-23, 18-22, 18-21, 18-20, 19-30, 19-29, 19-28, 19-27, 19-26, 19-25,19-24, 19-23, 19-22, 19-21, 19-20, 20-30, 20-29, 20-28, 20-27, 20-26,20-25, 20-24, 20-23, 20-22, 20-21, 21-30, 21-29, 21-28, 21-27, 21-26,21-25, 21-24, 21-23, or 21-22 base pairs in length. In certainembodiments, the duplex structure is 18 to 25 base pairs in length,e.g., 18-25, 18-24, 18-23, 18-22, 18-21, 18-20, 19-25, 19-24, 19-23,19-22, 19-21, 19-20, 20-25, 20-24, 20-23, 20-22, 20-21, 21-25, 21-24,21-23, 21-22, 22-25, 22-24, 22-23, 23-25, 23-24 or 24-25 base pairs inlength, for example, 19-21 basepairs in length. Ranges and lengthsintermediate to the above recited ranges and lengths are alsocontemplated to be part of the disclosure.

Similarly, the region of complementarity to the target sequence is 15 to30 nucleotides in length, e.g., 15-29, 15-28, 15-27, 15-26, 15-25,15-24, 15-23, 15-22, 15-21, 15-20, 15-19, 15-18, 15-17, 18-30, 18-29,18-28, 18-27, 18-26, 18-25, 18-24, 18-23, 18-22, 18-21, 18-20, 19-30,19-29, 19-28, 19-27, 19-26, 19-25, 19-24, 19-23, 19-22, 19-21, 19-20,20-30, 20-29, 20-28, 20-27, 20-26, 20-25, 20-24, 20-23, 20-22, 20-21,21-30, 21-29, 21-28, 21-27, 21-26, 21-25, 21-24, 21-23, or 21-22nucleotides in length, for example 19-23 nucleotides in length or 21-23nucleotides in length. Ranges and lengths intermediate to the aboverecited ranges and lengths are also contemplated to be part of thedisclosure.

In some embodiments, the duplex structure is 19 to 30 base pairs inlength. Similarly, the region of complementarity to the target sequenceis 19 to 30 nucleotides in length.

In some embodiments, the dsRNA is about 19 to about 23 nucleotides inlength, or about 25 to about 30 nucleotides in length. In general, thedsRNA is long enough to serve as a substrate for the Dicer enzyme. Forexample, it is well-known in the art that dsRNAs longer than about 21-23nucleotides in length may serve as substrates for Dicer. As theordinarily skilled person will also recognize, the region of an RNAtargeted for cleavage will most often be part of a larger RNA molecule,often an mRNA molecule. Where relevant, a “part” of an mRNA target is acontiguous sequence of an mRNA target of sufficient length to allow itto be a substrate for RNAi-directed cleavage (i.e., cleavage through aRISC pathway).

One of skill in the art will also recognize that the duplex region is aprimary functional portion of a dsRNA, e.g., a duplex region of about 19to about 30 base pairs, e.g., about 19-30, 19-29, 19-28, 19-27, 19-26,19-25, 19-24, 19-23, 19-22, 19-21, 19-20, 20-30, 20-29, 20-28, 20-27,20-26, 20-25, 20-24, 20-23, 20-22, 20-21, 21-30, 21-29, 21-28, 21-27,21-26, 21-25, 21-24, 21-23, or 21-22 base pairs. Thus, in oneembodiment, to the extent that it becomes processed to a functionalduplex, of e.g., 15-30 base pairs, that targets a desired RNA forcleavage, an RNA molecule or complex of RNA molecules having a duplexregion greater than 30 base pairs is a dsRNA. Thus, an ordinarilyskilled artisan will recognize that in one embodiment, a miRNA is adsRNA. In another embodiment, a dsRNA is not a naturally occurringmiRNA. In another embodiment, an iRNA agent useful to target complementfactor B gene expression is not generated in the target cell by cleavageof a larger dsRNA.

A dsRNA as described herein can further include one or moresingle-stranded nucleotide overhangs e.g., 1-4, 2-4, 1-3, 2-3, 1, 2, 3,or 4 nucleotides. dsRNAs having at least one nucleotide overhang canhave superior inhibitory properties relative to their blunt-endedcounterparts. A nucleotide overhang can comprise or consist of anucleotide/nucleoside analog, including a deoxynucleotide/nucleoside.The overhang(s) can be on the sense strand, the antisense strand, or anycombination thereof. Furthermore, the nucleotide(s) of an overhang canbe present on the 5′-end, 3′-end, or both ends of an antisense or sensestrand of a dsRNA.

A dsRNA can be synthesized by standard methods known in the art. Doublestranded RNAi compounds of the invention may be prepared using atwo-step procedure. First, the individual strands of the double strandedRNA molecule are prepared separately. Then, the component strands areannealed. The individual strands of the siRNA compound can be preparedusing solution-phase or solid-phase organic synthesis or both. Organicsynthesis offers the advantage that the oligonucleotide strandscomprising unnatural or modified nucleotides can be easily prepared.Similarly, single-stranded oligonucleotides of the invention can beprepared using solution-phase or solid-phase organic synthesis or both.

Regardless of the method of synthesis, the siRNA preparation can beprepared in a solution (e.g., an aqueous or organic solution) that isappropriate for formulation. For example, the siRNA preparation can beprecipitated and redissolved in pure double-distilled water, andlyophilized. The dried siRNA can then be resuspended in a solutionappropriate for the intended formulation process.

In an aspect, a dsRNA of the invention includes at least two nucleotidesequences, a sense sequence and an anti-sense sequence. The sense strandis selected from the group of sequences provided in any one of Tables2-3, and the corresponding antisense strand of the sense strand isselected from the group of sequences of any one of Tables 2-3. In thisaspect, one of the two sequences is complementary to the other of thetwo sequences, with one of the sequences being substantiallycomplementary to a sequence of an mRNA generated in the expression of acomplement factor B gene. As such, in this aspect, a dsRNA will includetwo oligonucleotides, where one oligonucleotide is described as thesense strand in any one of Tables 2-3, and the second oligonucleotide isdescribed as the corresponding antisense strand of the sense strand inany one of Tables 2-3.

In certain embodiments, the substantially complementary sequences of thedsRNA are contained on separate oligonucleotides. In other embodiments,the substantially complementary sequences of the dsRNA are contained ona single oligonucleotide.

It will be understood that, although the sequences in Table 2 are notdescribed as modified or conjugated sequences, the RNA of the iRNA ofthe invention e.g., a dsRNA of the invention, may comprise any one ofthe sequences set forth in any one of Tables 2-3 that is un-modified,un-conjugated, or modified or conjugated differently than describedtherein. In other words, the invention encompasses dsRNA of Tables 2-3which are un-modified, un-conjugated, modified, or conjugated, asdescribed herein.

The skilled person is well aware that dsRNAs having a duplex structureof about 20 to 23 base pairs, e.g., 21, base pairs have been hailed asparticularly effective in inducing RNA interference (Elbashir et al.,EMBO 2001, 20:6877-6888). However, others have found that shorter orlonger RNA duplex structures can also be effective (Chu and Rana (2007)RNA 14:1714-1719; Kim et al. (2005) Nat Biotech 23:222-226). In theembodiments described above, by virtue of the nature of theoligonucleotide sequences provided in any one of Tables 2-3, dsRNAsdescribed herein can include at least one strand of a length ofminimally 21 nucleotides. It can be reasonably expected that shorterduplexes having any one of the sequences in any one of Tables 2-3 minusonly a few nucleotides on one or both ends can be similarly effective ascompared to the dsRNAs described above. Hence, dsRNAs having a sequenceof at least 19, 20, or more contiguous nucleotides derived from any oneof the sequences of any one of Tables 2-3, and differing in theirability to inhibit the expression of a complement factor B gene by notmore than about 5, 10, 15, 20, 25, or 30% inhibition from a dsRNAcomprising the full sequence, are contemplated to be within the scope ofthe present invention.

In addition, the RNAs provided in Tables 2-3 identify a site(s) in acomplement factor B transcript that is susceptible to RISC-mediatedcleavage. As such, the present invention further features iRNAs thattarget within one of these sites. As used herein, an iRNA is said totarget within a particular site of an RNA transcript if the iRNApromotes cleavage of the transcript anywhere within that particularsite. Such an iRNA will generally include at least about 19 contiguousnucleotides from any one of the sequences provided in any one of Tables2-3 coupled to additional nucleotide sequences taken from the regioncontiguous to the selected sequence in a complement factor B gene.

An RNAi agent as described herein can contain one or more mismatches tothe target sequence. In one embodiment, an RNAi agent as describedherein contains no more than 3 mismatches (i.e., 3, 2, 1, or 0mismatches). In one embodiment, an RNAi agent as described hereincontains no more than 2 mismatches. In one embodiment, an RNAi agent asdescribed herein contains no more than 1 mismatch. In one embodiment, anRNAi agent as described herein contains 0 mismatches. In certainembodiments, if the antisense strand of the RNAi agent containsmismatches to the target sequence, the mismatch can optionally berestricted to be within the last 5 nucleotides from either the 5′- or3′-end of the region of complementarity. For example, in suchembodiments, for a 23 nucleotide RNAi agent, the strand which iscomplementary to a region of a CFB gene generally does not contain anymismatch within the central 13 nucleotides. The methods described hereinor methods known in the art can be used to determine whether an RNAiagent containing a mismatch to a target sequence is effective ininhibiting the expression of a CFB gene. Consideration of the efficacyof RNAi agents with mismatches in inhibiting expression of a CFB gene isimportant, especially if the particular region of complementarity in aCFB gene is known to have polymorphic sequence variation within thepopulation.

III. Modified iRNAs of the Invention

In certain embodiments, the RNA of the iRNA of the invention e.g., adsRNA, is un-modified, and does not comprise, e.g., chemicalmodifications or conjugations known in the art and described herein. Inother embodiments, the RNA of an iRNA of the invention, e.g., a dsRNA,is chemically modified to enhance stability or other beneficialcharacteristics. In certain embodiments of the invention, substantiallyall of the nucleotides of an iRNA of the invention are modified. Inother embodiments of the invention, all of the nucleotides of an iRNA orsubstantially all of the nucleotides of an iRNA are modified, i.e., notmore than 5, 4, 3, 2, or 1 unmodified nucleotides are present in astrand of the iRNA.

The nucleic acids featured in the invention can be synthesized ormodified by methods well established in the art, such as those describedin “Current protocols in nucleic acid chemistry,” Beaucage, S. L. et al.(Edrs.), John Wiley & Sons, Inc., New York, N.Y., USA, which is herebyincorporated herein by reference. Modifications include, for example,end modifications, e.g., 5′-end modifications (phosphorylation,conjugation, inverted linkages) or 3′-end modifications (conjugation,DNA nucleotides, inverted linkages, etc.); base modifications, e.g.,replacement with stabilizing bases, destabilizing bases, or bases thatbase pair with an expanded repertoire of partners, removal of bases(abasic nucleotides), or conjugated bases; sugar modifications (e.g., atthe 2′-position or 4′-position) or replacement of the sugar; or backbonemodifications, including modification or replacement of thephosphodiester linkages. Specific examples of iRNA compounds useful inthe embodiments described herein include, but are not limited to RNAscontaining modified backbones or no natural internucleoside linkages.RNAs having modified backbones include, among others, those that do nothave a phosphorus atom in the backbone. For the purposes of thisspecification, and as sometimes referenced in the art, modified RNAsthat do not have a phosphorus atom in their internucleoside backbone canalso be considered to be oligonucleosides. In some embodiments, amodified iRNA will have a phosphorus atom in its internucleosidebackbone.

Modified RNA backbones include, for example, phosphorothioates, chiralphosphorothioates, phosphorodithioates, phosphotriesters,aminoalkylphosphotriesters, methyl and other alkyl phosphonatesincluding 3′-alkylene phosphonates and chiral phosphonates,phosphinates, phosphoramidates including 3′-amino phosphoramidate andaminoalkylphosphoramidates, thionophosphoramidates,thionoalkylphosphonates, thionoalkylphosphotriesters, andboranophosphates having normal 3′-5′ linkages, 2′-5′-linked analogs ofthese, and those having inverted polarity wherein the adjacent pairs ofnucleoside units are linked 3′-5′ to 5′-3′ or 2′-5′ to 5′-2′. Varioussalts, mixed salts and free acid forms are also included. In someembodiments of the invention, the dsRNA agents of the invention are in afree acid form. In other embodiments of the invention, the dsRNA agentsof the invention are in a salt form. In one embodiment, the dsRNA agentsof the invention are in a sodium salt form. In certain embodiments, whenthe dsRNA agents of the invention are in the sodium salt form, sodiumions are present in the agent as counterions for substantially all ofthe phosphodiester or phosphorothioate groups present in the agent.Agents in which substantially all of the phosphodiester orphosphorothioate linkages have a sodium counterion include not more than5, 4, 3, 2, or 1 phosphodiester or phosphorothioate linkages without asodium counterion. In some embodiments, when the dsRNA agents of theinvention are in the sodium salt form, sodium ions are present in theagent as counterions for all of the phosphodiester or phosphorothioategroups present in the agent.

Representative U.S. patents that teach the preparation of the abovephosphorus-containing linkages include, but are not limited to, U.S.Pat. Nos. 3,687,808; 4,469,863; 4,476,301; 5,023,243; 5,177,195;5,188,897; 5,264,423; 5,276,019; 5,278,302; 5,286,717; 5,321,131;5,399,676; 5,405,939; 5,453,496; 5,455,233; 5,466,677; 5,476,925;5,519,126; 5,536,821; 5,541,316; 5,550,111; 5,563,253; 5,571,799;5,587,361; 5,625,050; 6,028,188; 6,124,445; 6,160,109; 6,169,170;6,172,209; 6,239,265; 6,277,603; 6,326,199; 6,346,614; 6,444,423;6,531,590; 6,534,639; 6,608,035; 6,683,167; 6,858,715; 6,867,294;6,878,805; 7,015,315; 7,041,816; 7,273,933; 7,321,029; and U.S. Pat.RE39464, the entire contents of each of which are hereby incorporatedherein by reference.

Modified RNA backbones that do not include a phosphorus atom thereinhave backbones that are formed by short chain alkyl or cycloalkylinternucleoside linkages, mixed heteroatoms and alkyl or cycloalkylinternucleoside linkages, or one or more short chain heteroatomic orheterocyclic internucleoside linkages. These include those havingmorpholino linkages (formed in part from the sugar portion of anucleoside); siloxane backbones; sulfide, sulfoxide and sulfonebackbones; formacetyl and thioformacetyl backbones; methylene formacetyland thioformacetyl backbones; alkene containing backbones; sulfamatebackbones; methyleneimino and methylenehydrazino backbones; sulfonateand sulfonamide backbones; amide backbones; and others having mixed N,O, S, and CH₂ component parts.

Representative U.S. patents that teach the preparation of the aboveoligonucleosides include, but are not limited to, U.S. Pat. Nos.5,034,506; 5,166,315; 5,185,444; 5,214,134; 5,216,141; 5,235,033;5,64,562; 5,264,564; 5,405,938; 5,434,257; 5,466,677; 5,470,967;5,489,677; 5,541,307; 5,561,225; 5,596,086; 5,602,240; 5,608,046;5,610,289; 5,618,704; 5,623,070; 5,663,312; 5,633,360; 5,677,437; and5,677,439, the entire contents of each of which are hereby incorporatedherein by reference.

Suitable RNA mimetics are contemplated for use in iRNAs provided herein,in which both the sugar and the internucleoside linkage, i.e., thebackbone, of the nucleotide units are replaced with novel groups. Thebase units are maintained for hybridization with an appropriate nucleicacid target compound. One such oligomeric compound in which an RNAmimetic that has been shown to have excellent hybridization propertiesis referred to as a peptide nucleic acid (PNA). In PNA compounds, thesugar backbone of an RNA is replaced with an amide containing backbone,in particular an aminoethylglycine backbone. The nucleobases areretained and are bound directly or indirectly to aza nitrogen atoms ofthe amide portion of the backbone. Representative US patents that teachthe preparation of PNA compounds include, but are not limited to, U.S.Pat. Nos. 5,539,082; 5,714,331; and 5,719,262, the entire contents ofeach of which are hereby incorporated herein by reference. AdditionalPNA compounds suitable for use in the iRNAs of the invention aredescribed in, for example, in Nielsen et al., Science, 1991, 254,1497-1500.

Some embodiments featured in the invention include RNAs withphosphorothioate backbones and oligonucleosides with heteroatombackbones, and in particular —CH₂—NH—CH₂—, —CH₂—N(CH₃)—O—CH₂— [known asa methylene (methylimino) or MMI backbone], —CH₂—O—N(CH₃)—CH₂—,—CH₂—N(CH₃)—N(CH₃)—CH₂— and —N(CH₃)—CH₂—CH₂— of the above-referencedU.S. Pat. No. 5,489,677, and the amide backbones of the above-referencedU.S. Pat. No. 5,602,240. In some embodiments, the RNAs featured hereinhave morpholino backbone structures of the above-referenced U.S. Pat.No. 5,034,506. The native phosphodiester backbone can be represented asO—P(O)(OH)—OCH₂—.

Modified RNAs can also contain one or more substituted sugar moieties.The iRNAs, e.g., dsRNAs, featured herein can include one of thefollowing at the 2′-position: OH; F; O-, S-, or N-alkyl; O-, S-, orN-alkenyl; O-, S- or N-alkynyl; or O-alkyl-O-alkyl, wherein the alkyl,alkenyl and alkynyl can be substituted or unsubstituted C₁ to C₁₀ alkylor C₂ to C₁₀ alkenyl and alkynyl. Exemplary suitable modificationsinclude O[(CH₂)_(n)O]_(m)CH₃, O(CH₂)·_(n)OCH₃, O(CH₂)_(n)NH₂,O(CH₂)_(n)CH₃, O(CH₂)_(n)ONH₂, and O(CH₂)_(n)ON[(CH₂)_(n)CH₃)]₂, where nand m are from 1 to about 10. In other embodiments, dsRNAs include oneof the following at the 2′ position: C₁ to C₁₀ lower alkyl, substitutedlower alkyl, alkaryl, aralkyl, O-alkaryl or O-aralkyl, SH, SCH₃, OCN,Cl, Br, CN, CF₃, OCF₃, SOCH₃, SO₂CH₃, ONO₂, NO₂, N₃, NH₂,heterocycloalkyl, heterocycloalkaryl, aminoalkylamino, polyalkylamino,substituted silyl, an RNA cleaving group, a reporter group, anintercalator, a group for improving the pharmacokinetic properties of aniRNA, or a group for improving the pharmacodynamic properties of aniRNA, and other substituents having similar properties. In someembodiments, the modification includes a 2′-methoxyethoxy(2′-O—CH₂CH₂OCH₃, also known as 2′-O-(2-methoxyethyl) or 2′-MOE) (Martinet al., Helv. Chim. Acta, 1995, 78:486-504) i.e., an alkoxy-alkoxygroup. Another exemplary modification is 2′-dimethylaminooxyethoxy,i.e., a O(CH₂)₂ON(CH₃)₂ group, also known as 2′-DMAOE, as described inexamples herein below, and 2′-dimethylaminoethoxyethoxy (also known inthe art as 2′-O-dimethylaminoethoxyethyl or 2′-DMAEOE), i.e.,2′-O—CH₂—O—CH₂—N(CH₂)₂. Further exemplary modifications include:5′-Me-2′-F nucleotides, 5′-Me-2′-OMe nucleotides,5′-Me-2′-deoxynucleotides, (both R and S isomers in these threefamilies); 2′-alkoxyalkyl; and 2′-NMA (N-methylacetamide).

Other modifications include 2′-methoxy (2′-OCH₃), 2′-aminopropoxy(2′-OCH₂CH₂CH₂NH₂) and 2′-fluoro (2′-F). Similar modifications can alsobe made at other positions on the RNA of an iRNA, particularly the 3′position of the sugar on the 3′ terminal nucleotide or in 2-5′ linkeddsRNAs and the 5′ position of 5′ terminal nucleotide. iRNAs can alsohave sugar mimetics such as cyclobutyl moieties in place of thepentofuranosyl sugar. Representative US patents that teach thepreparation of such modified sugar structures include, but are notlimited to, U.S. Pat. Nos. 4,981,957; 5,118,800; 5,319,080; 5,359,044;5,393,878; 5,446,137; 5,466,786; 5,514,785; 5,519,134; 5,567,811;5,576,427; 5,591,722; 5,597,909; 5,610,300; 5,627,053; 5,639,873;5,646,265; 5,658,873; 5,670,633; and 5,700,920, certain of which arecommonly owned with the instant application. The entire contents of eachof the foregoing are hereby incorporated herein by reference.

An iRNA can also include nucleobase (often referred to in the art simplyas “base”) modifications or substitutions. As used herein, “unmodified”or “natural” nucleobases include the purine bases adenine (A) andguanine (G), and the pyrimidine bases thymine (T), cytosine (C), anduracil (U). Modified nucleobases include other synthetic and naturalnucleobases such as deoxythimidine (dT), 5-methylcytosine (5-me-C),5-hydroxymethyl cytosine, xanthine, hypoxanthine, 2-aminoadenine,6-methyl and other alkyl derivatives of adenine and guanine, 2-propyland other alkyl derivatives of adenine and guanine, 2-thiouracil,2-thiothymine and 2-thiocytosine, 5-halouracil and cytosine, 5-propynyluracil and cytosine, 6-azo uracil, cytosine and thymine, 5-uracil(pseudouracil), 4-thiouracil, 8-halo, 8-amino, 8-thiol, 8-thioalkyl,8-hydroxyl anal other 8-substituted adenines and guanines, 5-halo,particularly 5-bromo, 5-trifluoromethyl and other 5-substituted uracilsand cytosines, 7-methylguanine and 7-methyladenine, 8-azaguanine and8-azaadenine, 7-deazaguanine and 7-daazaadenine and 3-deazaguanine and3-deazaadenine. Further nucleobases include those disclosed in U.S. Pat.No. 3,687,808, those disclosed in Modified Nucleosides in Biochemistry,Biotechnology and Medicine, Herdewijn, P. ed. Wiley-VCH, 2008; thosedisclosed in The Concise Encyclopedia Of Polymer Science AndEngineering, pages 858-859, Kroschwitz, J. L, ed. John Wiley & Sons,1990, these disclosed by Englisch et al., Angewandte Chemie,International Edition, 1991, 30, 613, and those disclosed by Sanghvi, YS., Chapter 15, dsRNA Research and Applications, pages 289-302, Crooke,S. T. and Lebleu, B., Ed., CRC Press, 1993. Certain of these nucleobasesare particularly useful for increasing the binding affinity of theoligomeric compounds featured in the invention. These include5-substituted pyrimidines, 6-azapyrimidines and N-2, N-6 and 0-6substituted purines, including 2-aminopropyladenine, 5-propynyluraciland 5-propynylcytosine. 5-methylcytosine substitutions have been shownto increase nucleic acid duplex stability by 0.6-1.2° C. (Sanghvi, Y.S., Crooke, S. T. and Lebleu, B., Eds., dsRNA Research and Applications,CRC Press, Boca Raton, 1993, pp. 276-278) and are exemplary basesubstitutions, even more particularly when combined with2′-O-methoxyethyl sugar modifications.

Representative U.S. patents that teach the preparation of certain of theabove noted modified nucleobases as well as other modified nucleobasesinclude, but are not limited to, the above noted U.S. Pat. Nos.3,687,808, 4,845,205; 5,130,30; 5,134,066; 5,175,273; 5,367,066;5,432,272; 5,457,187; 5,459,255; 5,484,908; 5,502,177; 5,525,711;5,552,540; 5,587,469; 5,594,121, 5,596,091; 5,614,617; 5,681,941;5,750,692; 6,015,886; 6,147,200; 6,166,197; 6,222,025; 6,235,887;6,380,368; 6,528,640; 6,639,062; 6,617,438; 7,045,610; 7,427,672; and7,495,088, the entire contents of each of which are hereby incorporatedherein by reference.

In some embodiments, an iRNA agent of the disclosure can also bemodified to include one or more bicyclic sugar moieties. A “bicyclicsugar” is a furanosyl ring modified by the bridging of two atoms. A“bicyclic sugar” is a furanosyl ring modified by a ring formed by thebridging of two carbons, whether adjacent or non-adjacent atoms. A“bicyclic nucleoside” (“BNA”) is a nucleoside having a sugar moietycomprising a bridge a ring formed by bridging connecting two carbons,whether adjacent or non-adjacent, atoms of the sugar ring, therebyforming a bicyclic ring system. In certain embodiments, the bridgeconnects the 4′-carbon and the 2′-carbon of the sugar ring, optionally,via the 2′-acyclic oxygen atoms. Thus, in some embodiments an agent ofthe invention may include one or more locked nucleic acids (LNA). Alocked nucleic acid is a nucleotide having a modified ribose moiety inwhich the ribose moiety comprises an extra bridge connecting the 2′ and4′ carbons. In other words, an LNA is a nucleotide comprising a bicyclicsugar moiety comprising a 4′-CH₂—O-2′ bridge. This structure effectively“locks” the ribose in the 3′-endo structural conformation. The additionof locked nucleic acids to siRNAs has been shown to increase siRNAstability in serum, and to reduce off-target effects (Elmen, J. et al.,(2005) Nucleic Acids Research 33(1):439-447; Mook, O R. et al., (2007)Mol Canc Ther 6(3):833-843; Grunweller, A. et al., (2003) Nucleic AcidsResearch 31(12):3185-3193). Examples of bicyclic nucleosides for use inthe polynucleotides of the invention include without limitationnucleosides comprising a bridge between the 4′ and the 2′ ribosyl ringatoms. In certain embodiments, the antisense polynucleotide agents ofthe invention include one or more bicyclic nucleosides comprising a 4′to 2′ bridge.

A locked nucleoside can be represented by the structure (omittingstereochemistry),

wherein B is a nucleobase or modified nucleobase and L is the linkinggroup that joins the 2′-carbon to the 4′-carbon of the ribose ring.

Examples of such 4′ to 2′ bridged bicyclic nucleosides, include but arenot limited to 4′-(CH₂)—O-2′ (LNA); 4′-(CH₂)—S-2′; 4′-(CH₂)₂—O-2′ (ENA);4′-CH(CH₃)—O-2′ (also referred to as “constrained ethyl” or “cEt”) and4′-CH(CH₂OCH₃)—O-2′ (and analogs thereof; see, e.g., U.S. Pat. No.7,399,845); 4′-C(CH₃)(CH₃)—O-2′ (and analogs thereof; see e.g., U.S.Pat. No. 8,278,283); 4′-CH₂—N(OCH₃)-2′ (and analogs thereof; see e.g.,U.S. Pat. No. 8,278,425); 4′-CH₂—O—N(CH₃)-2′ (see, e.g., U.S. PatentPublication No. 2004/0171570); 4′-CH₂—N(R)—O-2′, wherein R is H, C1-C12alkyl, or a nitrogen protecting group (see, e.g., U.S. Pat. No.7,427,672); 4′-CH₂—C(H)(CH₃)-2′ (see, e.g., Chattopadhyaya et al., J.Org. Chem., 2009, 74, 118-134); and 4′-CH₂—C(═CH₂)-2′ (and analogsthereof; see, e.g., U.S. Pat. No. 8,278,426). The entire contents ofeach of the foregoing are hereby incorporated herein by reference.

Additional representative U.S. patents and U.S. Patent Publications thatteach the preparation of locked nucleic acid nucleotides include, butare not limited to, the following: U.S. Pat. Nos. 6,268,490; 6,525,191;6,670,461; 6,770,748; 6,794,499; 6,998,484; 7,053,207; 7,034,133;7,084,125; 7,399,845; 7,427,672; 7,569,686; 7,741,457; 8,022,193;8,030,467; 8,278,425; 8,278,426; 8,278,283; US 2008/0039618; and US2009/0012281, the entire contents of each of which are herebyincorporated herein by reference.

Any of the foregoing bicyclic nucleosides can be prepared having one ormore stereochemical sugar configurations including for exampleα-L-ribofuranose and β-D-ribofuranose (see WO 99/14226).

An iRNA agent of the disclosure can also be modified to include one ormore constrained ethyl nucleotides. As used herein, a “constrained ethylnucleotide” or “cEt” is a locked nucleic acid comprising a bicyclicsugar moiety comprising a 4′-CH(CH₃)—O-2′ bridge (i.e., L in thepreceding structure). In one embodiment, a constrained ethyl nucleotideis in the S conformation referred to herein as “S-cEt.”

An iRNA of the invention may also include one or more “conformationallyrestricted nucleotides” (“CRN”). CRN are nucleotide analogs with alinker connecting the C2′ and C4′ carbons of ribose or the CFB and —C5′carbons of ribose. CRN lock the ribose ring into a stable conformationand increase the hybridization affinity to mRNA. The linker is ofsufficient length to place the oxygen in an optimal position forstability and affinity resulting in less ribose ring puckering.

Representative publications that teach the preparation of certain of theabove noted CRN include, but are not limited to, US2013/0190383; andWO2013/036868, the entire contents of each of which are herebyincorporated herein by reference.

In some embodiments, an iRNA of the invention comprises one or moremonomers that are UNA (unlocked nucleic acid) nucleotides. UNA isunlocked acyclic nucleic acid, wherein any of the bonds of the sugar hasbeen removed, forming an unlocked “sugar” residue. In one example, UNAalso encompasses monomer with bonds between C1′-C4′ have been removed(i.e. the covalent carbon-oxygen-carbon bond between the C1′ and C4′carbons). In another example, the C2′-C3′ bond (i.e. the covalentcarbon-carbon bond between the C2′ and C3′ carbons) of the sugar hasbeen removed (see Nuc. Acids Symp. Series, 52, 133-134 (2008) andFluiter et al., Mol. Biosyst., 2009, 10, 1039 hereby incorporated byreference).

Representative U.S. publications that teach the preparation of UNAinclude, but are not limited to, U.S. Pat. No. 8,314,227; andUS2013/0096289; US2013/0011922; and US2011/0313020, the entire contentsof each of which are hereby incorporated herein by reference.

Potentially stabilizing modifications to the ends of RNA molecules caninclude N-(acetylaminocaproyl)-4-hydroxyprolinol (Hyp-C6-NHAc),N-(caproyl-4-hydroxyprolinol (Hyp-C6), N-(acetyl-4-hydroxyprolinol(Hyp-NHAc), thymidine-2′-O-deoxythymidine (ether),N-(aminocaproyl)-4-hydroxyprolinol (Hyp-C6-amino),2-docosanoyl-uridine-3′-phosphate, inverted 2′-deoxy-modifiedribonucleotide, such as inverted dT(idT), inverted dA (idA), andinverted abasic 2′-deoxyribonucleotide (iAb) and others. Disclosure ofthis modification can be found in WO 2011/005861.

In one example, the 3′ or 5′ terminal end of a oligonucleotide is linkedto an inverted 2′-deoxy-modified ribonucleotide, such as inverteddT(idT), inverted dA (idA), or a inverted abasic 2′-deoxyribonucleotide(iAb). In one particular example, the inverted 2′-deoxy-modifiedribonucleotide is linked to the 3′end of an oligonucleotide, such as the3′-end of a sense strand described herein, where the linking is via a3′-3′ phosphodiester linkage or a 3′-3′-phosphorothioate linkage.

In another example, the 3′-end of a sense strand is linked via a3′-3′-phosphorothioate linkage to an inverted abasic ribonucleotide(iAb). In another example, the 3′-end of a sense strand is linked via a3′-3′-phosphorothioate linkage to an inverted dA (idA).

In one particular example, the inverted 2′-deoxy-modified ribonucleotideis linked to the 3′end of an oligonucleotide, such as the 3′-end of asense strand described herein, where the linking is via a 3′-3′phosphodiester linkage or a 3′-3′-phosphorothioate linkage.

In another example, the 3′-terminal nucleotides of a sense strand is aninverted dA (idA) and is linked to the preceding nucleotide via a3′-3′-linkage (e.g., 3′-3′-phosphorothioate linkage).

Other modifications of the nucleotides of an iRNA of the inventioninclude a 5′ phosphate or 5′ phosphate mimic, e.g., a 5′-terminalphosphate or phosphate mimic on the antisense strand of an iRNA.Suitable phosphate mimics are disclosed in, for example US2012/0157511,the entire contents of which are incorporated herein by reference.

A. Modified iRNAs Comprising Motifs of the Invention

In certain aspects of the invention, the double stranded RNA agents ofthe invention include agents with chemical modifications as disclosed,for example, in WO2013/075035, the entire contents of each of which areincorporated herein by reference. As shown herein and in WO2013/075035,one or more motifs of three identical modifications on three consecutivenucleotides may be introduced into a sense strand or antisense strand ofa dsRNAi agent, particularly at or near the cleavage site. In someembodiments, the sense strand and antisense strand of the dsRNAi agentmay otherwise be completely modified. The introduction of these motifsinterrupts the modification pattern, if present, of the sense orantisense strand. The dsRNAi agent may be optionally conjugated with aGalNAc derivative ligand, for instance on the sense strand.

More specifically, when the sense strand and antisense strand of thedouble stranded RNA agent are completely modified to have one or moremotifs of three identical modifications on three consecutive nucleotidesat or near the cleavage site of at least one strand of a dsRNAi agent,the gene silencing activity of the dsRNAi agent was observed.

Accordingly, the invention provides double stranded RNA agents capableof inhibiting the expression of a target gene (i.e., CFB gene) in vivo.The RNAi agent comprises a sense strand and an antisense strand. Eachstrand of the RNAi agent may be, for example, 17-30 nucleotides inlength, 25-30 nucleotides in length, 27-30 nucleotides in length, 19-25nucleotides in length, 19-23 nucleotides in length, 19-21 nucleotides inlength, 21-25 nucleotides in length, or 21-23 nucleotides in length.

The sense strand and antisense strand typically form a duplex doublestranded RNA (“dsRNA”), also referred to herein as “dsRNAi agent.” Theduplex region of a dsRNAi agent may be, for example, the duplex regioncan be 27-30 nucleotide pairs in length, 19-25 nucleotide pairs inlength, 19-23 nucleotide pairs in length, 19-21 nucleotide pairs inlength, 21-25 nucleotide pairs in length, or 21-23 nucleotide pairs inlength. In another example, the duplex region is selected from 19, 20,21, 22, 23, 24, 25, 26, and 27 nucleotides in length.

In certain embodiments, the dsRNAi agent may contain one or moreoverhang regions or capping groups at the 3′-end, 5′-end, or both endsof one or both strands. The overhang can be, independently, 1-6nucleotides in length, for instance 2-6 nucleotides in length, 1-5nucleotides in length, 2-5 nucleotides in length, 1-4 nucleotides inlength, 2-4 nucleotides in length, 1-3 nucleotides in length, 2-3nucleotides in length, or 1-2 nucleotides in length. In certainembodiments, the overhang regions can include extended overhang regionsas provided above. The overhangs can be the result of one strand beinglonger than the other, or the result of two strands of the same lengthbeing staggered. The overhang can form a mismatch with the target mRNAor it can be complementary to the gene sequences being targeted or canbe another sequence. The first and second strands can also be joined,e.g., by additional bases to form a hairpin, or by other non-baselinkers.

In certain embodiments, the nucleotides in the overhang region of thedsRNAi agent can each independently be a modified or unmodifiednucleotide including, but no limited to 2′-sugar modified, such as,2′-F, 2′-O-methyl, thymidine (T), 2′-O-methoxyethyl-5-methyluridine(Teo), 2′-O-methoxyethyladenosine (Aeo),2′-O-methoxyethyl-5-methylcytidine (m5Ceo), and any combinationsthereof.

For example, TT can be an overhang sequence for either end on eitherstrand. The overhang can form a mismatch with the target mRNA or it canbe complementary to the gene sequences being targeted or can be anothersequence.

The 5′- or 3′-overhangs at the sense strand, antisense strand, or bothstrands of the dsRNAi agent may be phosphorylated. In some embodiments,the overhang region(s) contains two nucleotides having aphosphorothioate between the two nucleotides, where the two nucleotidescan be the same or different. In some embodiments, the overhang ispresent at the 3′-end of the sense strand, antisense strand, or bothstrands. In some embodiments, this 3′-overhang is present in theantisense strand. In some embodiments, this 3′-overhang is present inthe sense strand.

The RNAi agent may contain only a single overhang, which can strengthenthe interference activity of the RNAi, without affecting its overallstability. For example, the single-stranded overhang may be located atthe 3′-end of the sense strand or, alternatively, at the 3′-end of theantisense strand. The RNAi may also have a blunt end, located at the5′-end of the antisense strand (i.e., the 3′-end of the sense strand) orvice versa. Generally, the antisense strand of the dsRNAi agent has anucleotide overhang at the 3′-end, and the 5′-end is blunt. While notwishing to be bound by theory, the asymmetric blunt end at the 5′-end ofthe antisense strand and 3′-end overhang of the antisense strand favorthe guide strand loading into RISC process.

In certain embodiments, the dsRNAi agent is a double blunt-ended of 19nucleotides in length, wherein the sense strand contains at least onemotif of three 2′-F modifications on three consecutive nucleotides atpositions 7, 8, and 9 from the 5′end. The antisense strand contains atleast one motif of three 2′-O-methyl modifications on three consecutivenucleotides at positions 11, 12, and 13 from the 5′end.

In other embodiments, the dsRNAi agent is a double blunt-ended of 20nucleotides in length, wherein the sense strand contains at least onemotif of three 2′-F modifications on three consecutive nucleotides atpositions 8, 9, and 10 from the 5′end. The antisense strand contains atleast one motif of three 2′-O-methyl modifications on three consecutivenucleotides at positions 11, 12, and 13 from the 5′end.

In yet other embodiments, the dsRNAi agent is a double blunt-ended of 21nucleotides in length, wherein the sense strand contains at least onemotif of three 2′-F modifications on three consecutive nucleotides atpositions 9, 10, and 11 from the 5′end. The antisense strand contains atleast one motif of three 2′-O-methyl modifications on three consecutivenucleotides at positions 11, 12, and 13 from the 5′end.

In certain embodiments, the dsRNAi agent comprises a 21 nucleotide sensestrand and a 23 nucleotide antisense strand, wherein the sense strandcontains at least one motif of three 2′-F modifications on threeconsecutive nucleotides at positions 9, 10, and 11 from the 5′ end; theantisense strand contains at least one motif of three 2′-O-methylmodifications on three consecutive nucleotides at positions 11, 12, and13 from the 5′end, wherein one end of the RNAi agent is blunt, while theother end comprises a two nucleotide overhang. in one embodiment, thetwo nucleotide overhang is at the 3′-end of the antisense strand.

When the two nucleotide overhang is at the 3′-end of the antisensestrand, there may be two phosphorothioate internucleotide linkagesbetween the terminal three nucleotides, wherein two of the threenucleotides are the overhang nucleotides, and the third nucleotide is apaired nucleotide next to the overhang nucleotide. In one embodiment,the RNAi agent additionally has two phosphorothioate internucleotidelinkages between the terminal three nucleotides at both the 5′-end ofthe sense strand and at the 5′-end of the antisense strand. In certainembodiments, every nucleotide in the sense strand and the antisensestrand of the dsRNAi agent, including the nucleotides that are part ofthe motifs are modified nucleotides. In certain embodiments each residueis independently modified with a 2′-O-methyl or 2′-fluoro, e.g., in analternating motif. Optionally, the dsRNAi agent further comprises aligand (such as, GalNAc).

In certain embodiments, the dsRNAi agent comprises a sense and anantisense strand, wherein the sense strand is 25-30 nucleotide residuesin length, wherein starting from the 5′ terminal nucleotide (position 1)positions 1 to 23 of the first strand comprise at least 8ribonucleotides; the antisense strand is 36-66 nucleotide residues inlength and, starting from the 3′ terminal nucleotide, comprises at least8 ribonucleotides in the positions paired with positions 1-23 of sensestrand to form a duplex; wherein at least the 3′ terminal nucleotide ofantisense strand is unpaired with sense strand, and up to 6 consecutive3′ terminal nucleotides are unpaired with sense strand, thereby forminga 3′ single stranded overhang of 1-6 nucleotides; wherein the 5′terminus of antisense strand comprises from 10-30 consecutivenucleotides which are unpaired with sense strand, thereby forming a10-30 nucleotide single stranded 5′ overhang; wherein at least the sensestrand 5′ terminal and 3′ terminal nucleotides are base paired withnucleotides of antisense strand when sense and antisense strands arealigned for maximum complementarity, thereby forming a substantiallyduplexed region between sense and antisense strands; and antisensestrand is sufficiently complementary to a target RNA along at least 19ribonucleotides of antisense strand length to reduce target geneexpression when the double stranded nucleic acid is introduced into amammalian cell; and wherein the sense strand contains at least one motifof three 2′-F modifications on three consecutive nucleotides, where atleast one of the motifs occurs at or near the cleavage site. Theantisense strand contains at least one motif of three 2′-O-methylmodifications on three consecutive nucleotides at or near the cleavagesite.

In certain embodiments, the dsRNAi agent comprises sense and antisensestrands, wherein the dsRNAi agent comprises a first strand having alength which is at least 25 and at most 29 nucleotides and a secondstrand having a length which is at most 30 nucleotides with at least onemotif of three 2′-O-methyl modifications on three consecutivenucleotides at position 11, 12, and 13 from the 5′ end; wherein the 3′end of the first strand and the 5′ end of the second strand form a bluntend and the second strand is 1-4 nucleotides longer at its 3′ end thanthe first strand, wherein the duplex region which is at least 25nucleotides in length, and the second strand is sufficientlycomplementary to a target mRNA along at least 19 nucleotide of thesecond strand length to reduce target gene expression when the RNAiagent is introduced into a mammalian cell, and wherein Dicer cleavage ofthe dsRNAi agent results in an siRNA comprising the 3′-end of the secondstrand, thereby reducing expression of the target gene in the mammal.Optionally, the dsRNAi agent further comprises a ligand.

In certain embodiments, the sense strand of the dsRNAi agent contains atleast one motif of three identical modifications on three consecutivenucleotides, where one of the motifs occurs at the cleavage site in thesense strand.

In certain embodiments, the antisense strand of the dsRNAi agent canalso contain at least one motif of three identical modifications onthree consecutive nucleotides, where one of the motifs occurs at or nearthe cleavage site in the antisense strand.

For a dsRNAi agent having a duplex region of 19-23 nucleotides inlength, the cleavage site of the antisense strand is typically aroundthe 10, 11, and 12 positions from the 5′-end. Thus the motifs of threeidentical modifications may occur at the 9, 10, and 11 positions; the10, 11, and 12 positions; the 11, 12, and 13 positions; the 12, 13, and14 positions; or the 13, 14, and 15 positions of the antisense strand,the count starting from the first nucleotide from the 5′-end of theantisense strand, or, the count starting from the first pairednucleotide within the duplex region from the 5′-end of the antisensestrand. The cleavage site in the antisense strand may also changeaccording to the length of the duplex region of the dsRNAi agent fromthe 5′-end.

The sense strand of the dsRNAi agent may contain at least one motif ofthree identical modifications on three consecutive nucleotides at thecleavage site of the strand; and the antisense strand may have at leastone motif of three identical modifications on three consecutivenucleotides at or near the cleavage site of the strand. When the sensestrand and the antisense strand form a dsRNA duplex, the sense strandand the antisense strand can be so aligned that one motif of the threenucleotides on the sense strand and one motif of the three nucleotideson the antisense strand have at least one nucleotide overlap, i.e., atleast one of the three nucleotides of the motif in the sense strandforms a base pair with at least one of the three nucleotides of themotif in the antisense strand. Alternatively, at least two nucleotidesmay overlap, or all three nucleotides may overlap.

In some embodiments, the sense strand of the dsRNAi agent may containmore than one motif of three identical modifications on threeconsecutive nucleotides. The first motif may occur at or near thecleavage site of the strand and the other motifs may be a wingmodification. The term “wing modification” herein refers to a motifoccurring at another portion of the strand that is separated from themotif at or near the cleavage site of the same strand. The wingmodification is either adjacent to the first motif or is separated by atleast one or more nucleotides. When the motifs are immediately adjacentto each other then the chemistries of the motifs are distinct from eachother, and when the motifs are separated by one or more nucleotide thanthe chemistries can be the same or different. Two or more wingmodifications may be present. For instance, when two wing modificationsare present, each wing modification may occur at one end relative to thefirst motif which is at or near cleavage site or on either side of thelead motif.

Like the sense strand, the antisense strand of the dsRNAi agent maycontain more than one motifs of three identical modifications on threeconsecutive nucleotides, with at least one of the motifs occurring at ornear the cleavage site of the strand. This antisense strand may alsocontain one or more wing modifications in an alignment similar to thewing modifications that may be present on the sense strand.

In some embodiments, the wing modification on the sense strand orantisense strand of the dsRNAi agent typically does not include thefirst one or two terminal nucleotides at the 3′-end, 5′-end, or bothends of the strand.

In other embodiments, the wing modification on the sense strand orantisense strand of the dsRNAi agent typically does not include thefirst one or two paired nucleotides within the duplex region at the3′-end, 5′-end, or both ends of the strand.

When the sense strand and the antisense strand of the dsRNAi agent eachcontain at least one wing modification, the wing modifications may fallon the same end of the duplex region, and have an overlap of one, two,or three nucleotides.

When the sense strand and the antisense strand of the dsRNAi agent eachcontain at least two wing modifications, the sense strand and theantisense strand can be so aligned that two modifications each from onestrand fall on one end of the duplex region, having an overlap of one,two, or three nucleotides; two modifications each from one strand fallon the other end of the duplex region, having an overlap of one, two orthree nucleotides; two modifications one strand fall on each side of thelead motif, having an overlap of one, two or three nucleotides in theduplex region.

In some embodiments, every nucleotide in the sense strand and antisensestrand of the dsRNAi agent, including the nucleotides that are part ofthe motifs, may be modified. Each nucleotide may be modified with thesame or different modification which can include one or more alterationof one or both of the non-linking phosphate oxygens or of one or more ofthe linking phosphate oxygens; alteration of a constituent of the ribosesugar, e.g., of the 2′-hydroxyl on the ribose sugar; wholesalereplacement of the phosphate moiety with “dephospho” linkers;modification or replacement of a naturally occurring base; andreplacement or modification of the ribose-phosphate backbone.

As nucleic acids are polymers of subunits, many of the modificationsoccur at a position which is repeated within a nucleic acid, e.g., amodification of a base, or a phosphate moiety, or a non-linking O of aphosphate moiety. In some cases the modification will occur at all ofthe subject positions in the nucleic acid but in many cases it will not.By way of example, a modification may only occur at a 3′- or 5′ terminalposition, may only occur in a terminal region, e.g., at a position on aterminal nucleotide or in the last 2, 3, 4, 5, or 10 nucleotides of astrand. A modification may occur in a double strand region, a singlestrand region, or in both. A modification may occur only in the doublestrand region of an RNA or may only occur in a single strand region of aRNA. For example, a phosphorothioate modification at a non-linking Oposition may only occur at one or both termini, may only occur in aterminal region, e.g., at a position on a terminal nucleotide or in thelast 2, 3, 4, 5, or 10 nucleotides of a strand, or may occur in doublestrand and single strand regions, particularly at termini. The 5′-end orends can be phosphorylated.

It may be possible, e.g., to enhance stability, to include particularbases in overhangs, or to include modified nucleotides or nucleotidesurrogates, in single strand overhangs, e.g., in a 5′- or 3′-overhang,or in both. For example, it can be desirable to include purinenucleotides in overhangs. In some embodiments all or some of the basesin a 3′- or 5′-overhang may be modified, e.g., with a modificationdescribed herein. Modifications can include, e.g., the use ofmodifications at the 2′ position of the ribose sugar with modificationsthat are known in the art, e.g., the use of deoxyribonucleotides,2′-deoxy-2′-fluoro (2′-F) or 2′-O-methyl modified instead of theribosugar of the nucleobase, and modifications in the phosphate group,e.g., phosphorothioate modifications. Overhangs need not be homologouswith the target sequence.

In some embodiments, each residue of the sense strand and antisensestrand is independently modified with LNA, CRN, cET, UNA, HNA, CeNA,2′-methoxyethyl, 2′-O-methyl, 2′-O-allyl, 2′-C-allyl, 2′-deoxy,2′-hydroxyl, or 2′-fluoro. The strands can contain more than onemodification. In one embodiment, each residue of the sense strand andantisense strand is independently modified with 2′-O-methyl or2′-fluoro.

At least two different modifications are typically present on the sensestrand and antisense strand. Those two modifications may be the2′-O-methyl or 2′-fluoro modifications, or others.

In certain embodiments, the N_(a) or N_(b) comprise modifications of analternating pattern. The term “alternating motif” as used herein refersto a motif having one or more modifications, each modification occurringon alternating nucleotides of one strand. The alternating nucleotide mayrefer to one per every other nucleotide or one per every threenucleotides, or a similar pattern. For example, if A, B and C eachrepresent one type of modification to the nucleotide, the alternatingmotif can be “ABABABABABAB . . . ,” “AABBAABBAABB . . . ,” “AABAABAABAAB. . . ,” “AAABAAABAAAB . . . ,” “AAABBBAAABBB . . . ,” or “ABCABCABCABC. . . ,” etc.

The type of modifications contained in the alternating motif may be thesame or different. For example, if A, B, C, D each represent one type ofmodification on the nucleotide, the alternating pattern, i.e.,modifications on every other nucleotide, may be the same, but each ofthe sense strand or antisense strand can be selected from severalpossibilities of modifications within the alternating motif such as“ABABAB . . . ”, “ACACAC . . . ” “BDBDBD . . . ” or “CDCDCD . . . ,”etc.

In some embodiments, the dsRNAi agent of the invention comprises themodification pattern for the alternating motif on the sense strandrelative to the modification pattern for the alternating motif on theantisense strand is shifted. The shift may be such that the modifiedgroup of nucleotides of the sense strand corresponds to a differentlymodified group of nucleotides of the antisense strand and vice versa.For example, the sense strand when paired with the antisense strand inthe dsRNA duplex, the alternating motif in the sense strand may startwith “ABABAB” from 5′ to 3′ of the strand and the alternating motif inthe antisense strand may start with “BABABA” from 5′ to 3′ of the strandwithin the duplex region. As another example, the alternating motif inthe sense strand may start with “AABBAABB” from 5′ to 3′ of the strandand the alternating motif in the antisense strand may start with“BBAABBAA” from 5′ to 3′ of the strand within the duplex region, so thatthere is a complete or partial shift of the modification patternsbetween the sense strand and the antisense strand.

In one particular example, the alternating motif in the sense strand is“ABABAB” from 5′ 3′ of the strand, where each A is an unmodifiedribonucleotide and each B is a 2′-Omethyl modified nucleotide.

In one particular example, the alternating motif in the sense strand is“ABABAB” from 5′ 3′ of the strand, where each A is an 2′-deoxy-2′-fluoromodified nucleotide and each B is a 2′-Omethyl modified nucleotide.

In another particular example, the alternating motif in the antisensestrand is “BABABA” from 3′-5′ of the strand, where each A is a2′-deoxy-2′-fluoro modified nucleotide and each B is a 2′-Omethylmodified nucleotide.

In one particular example, the alternating motif in the sense strand is“ABABAB” from 5′ 3′ of the strand and the alternating motif in theantisense strand is “BABABA” from 3′-5′ of the strand, where each A isan unmodified ribonucleotide and each B is a 2′-Omethyl modifiednucleotide.

In one particular example, the alternating motif in the sense strand is“ABABAB” from 5′ 3′ of the strand and the alternating motif in theantisense strand is “BABABA” from 3′-5′ of the strand, where each A is a2′-deoxy-2′-fluoro modified nucleotide and each B is a 2′-Omethylmodified nucleotide.

In some embodiments, the dsRNAi agent comprises the pattern of thealternating motif of 2′-O-methyl modification and 2′-F modification onthe sense strand initially has a shift relative to the pattern of thealternating motif of 2′-O-methyl modification and 2′-F modification onthe antisense strand initially, i.e., the 2′-O-methyl modifiednucleotide on the sense strand base pairs with a 2′-F modifiednucleotide on the antisense strand and vice versa. The 1 position of thesense strand may start with the 2′-F modification, and the 1 position ofthe antisense strand may start with the 2′-O-methyl modification.

The introduction of one or more motifs of three identical modificationson three consecutive nucleotides to the sense strand or antisense strandinterrupts the initial modification pattern present in the sense strandor antisense strand. This interruption of the modification pattern ofthe sense or antisense strand by introducing one or more motifs of threeidentical modifications on three consecutive nucleotides to the sense orantisense strand may enhance the gene silencing activity against thetarget gene.

In some embodiments, when the motif of three identical modifications onthree consecutive nucleotides is introduced to any of the strands, themodification of the nucleotide next to the motif is a differentmodification than the modification of the motif. For example, theportion of the sequence containing the motif is “ . . . N_(a)YYYN_(b) .. . ,” where “Y” represents the modification of the motif of threeidentical modifications on three consecutive nucleotide, and “N_(a)” and“N_(b)” represent a modification to the nucleotide next to the motif“YYY” that is different than the modification of Y, and where N_(a) andN_(b) can be the same or different modifications. Alternatively, N_(a)or N_(b) may be present or absent when there is a wing modificationpresent.

The iRNA may further comprise at least one phosphorothioate ormethylphosphonate internucleotide linkage. The phosphorothioate ormethylphosphonate internucleotide linkage modification may occur on anynucleotide of the sense strand, antisense strand, or both strands in anyposition of the strand. For instance, the internucleotide linkagemodification may occur on every nucleotide on the sense strand orantisense strand; each internucleotide linkage modification may occur inan alternating pattern on the sense strand or antisense strand; or thesense strand or antisense strand may contain both internucleotidelinkage modifications in an alternating pattern. The alternating patternof the internucleotide linkage modification on the sense strand may bethe same or different from the antisense strand, and the alternatingpattern of the internucleotide linkage modification on the sense strandmay have a shift relative to the alternating pattern of theinternucleotide linkage modification on the antisense strand. In oneembodiment, a double-stranded RNAi agent comprises 6-8 phosphorothioateinternucleotide linkages. In some embodiments, the antisense strandcomprises two phosphorothioate internucleotide linkages at the 5′-endand two phosphorothioate internucleotide linkages at the 3′-end, and thesense strand comprises at least two phosphorothioate internucleotidelinkages at either the 5′-end or the 3′-end.

In some embodiments, the dsRNAi agent comprises a phosphorothioate ormethylphosphonate internucleotide linkage modification in the overhangregion. For example, the overhang region may contain two nucleotideshaving a phosphorothioate or methylphosphonate internucleotide linkagebetween the two nucleotides. Internucleotide linkage modifications alsomay be made to link the overhang nucleotides with the terminal pairednucleotides within the duplex region. For example, at least 2, 3, 4, orall the overhang nucleotides may be linked through phosphorothioate ormethylphosphonate internucleotide linkage, and optionally, there may beadditional phosphorothioate or methylphosphonate internucleotidelinkages linking the overhang nucleotide with a paired nucleotide thatis next to the overhang nucleotide. For instance, there may be at leasttwo phosphorothioate internucleotide linkages between the terminal threenucleotides, in which two of the three nucleotides are overhangnucleotides, and the third is a paired nucleotide next to the overhangnucleotide. These terminal three nucleotides may be at the 3′-end of theantisense strand, the 3′-end of the sense strand, the 5′-end of theantisense strand, or the 5′end of the antisense strand.

In some embodiments, the 2-nucleotide overhang is at the 3′-end of theantisense strand, and there are two phosphorothioate internucleotidelinkages between the terminal three nucleotides, wherein two of thethree nucleotides are the overhang nucleotides, and the third nucleotideis a paired nucleotide next to the overhang nucleotide. Optionally, thedsRNAi agent may additionally have two phosphorothioate internucleotidelinkages between the terminal three nucleotides at both the 5′-end ofthe sense strand and at the 5′-end of the antisense strand.

In one embodiment, the dsRNAi agent comprises mismatch(es) with thetarget, within the duplex, or combinations thereof. The mismatch mayoccur in the overhang region or the duplex region. The base pair may beranked on the basis of their propensity to promote dissociation ormelting (e.g., on the free energy of association or dissociation of aparticular pairing, the simplest approach is to examine the pairs on anindividual pair basis, though next neighbor or similar analysis can alsobe used). In terms of promoting dissociation: A:U is preferred over G:C;G:U is preferred over G:C; and I:C is preferred over G:C (I=inosine).Mismatches, e.g., non-canonical or other than canonical pairings (asdescribed elsewhere herein) are preferred over canonical (A:T, A:U, G:C)pairings; and pairings which include a universal base are preferred overcanonical pairings.

In certain embodiments, the dsRNAi agent comprises at least one of thefirst 1, 2, 3, 4, or 5 base pairs within the duplex regions from the5′-end of the antisense strand independently selected from the group of:A:U, G:U, I:C, and mismatched pairs, e.g., non-canonical or other thancanonical pairings or pairings which include a universal base, topromote the dissociation of the antisense strand at the 5′-end of theduplex.

In certain embodiments, the nucleotide at the 1 position within theduplex region from the 5′-end in the antisense strand is selected fromA, dA, dU, U, and dT. Alternatively, at least one of the first 1, 2, or3 base pair within the duplex region from the 5′-end of the antisensestrand is an AU base pair. For example, the first base pair within theduplex region from the 5′-end of the antisense strand is an AU basepair.

In other embodiments, the nucleotide at the 3′-end of the sense strandis deoxythimidine (dT) or the nucleotide at the 3′-end of the antisensestrand is deoxythimidine (dT). For example, there is a short sequence ofdeoxythimidine nucleotides, for example, two dT nucleotides on the3′-end of the sense, antisense strand, or both strands.

In certain embodiments, the sense strand sequence may be represented byformula (I):

5′n_(p)-N_(a)-(XXX)_(i)-N_(b)-YYY-N_(b)-(ZZZ)_(j)N_(a)-n_(q)3′  (I)

wherein:

i and j are each independently 0 or 1;

p and q are each independently 0-6;

each N_(a) independently represents an oligonucleotide sequencecomprising 0-25 modified nucleotides, each sequence comprising at leasttwo differently modified nucleotides;

each N_(b) independently represents an oligonucleotide sequencecomprising 0-10 modified nucleotides;

each n_(p) and n_(q) independently represent an overhang nucleotide;

wherein Nb and Y do not have the same modification; and

XXX, YYY, and ZZZ each independently represent one motif of threeidentical modifications on three consecutive nucleotides. In oneembodiment, YYY is all 2′-F modified nucleotides.

In some embodiments, the N_(a) or N_(b) comprises modifications ofalternating pattern.

In some embodiments, the YYY motif occurs at or near the cleavage siteof the sense strand. For example, when the dsRNAi agent has a duplexregion of 17-23 nucleotides in length, the YYY motif can occur at or thevicinity of the cleavage site (e.g.: can occur at positions 6, 7, 8; 7,8, 9; 8, 9, 10; 9, 10, 11; 10, 11, 12; or 11, 12, 13) of the sensestrand, the count starting from the first nucleotide, from the 5′-end;or optionally, the count starting at the first paired nucleotide withinthe duplex region, from the 5′-end.

In one embodiment, i is 1 and j is 0, or i is 0 and j is 1, or both iand j are 1. The sense strand can therefore be represented by thefollowing formulas:

5′n_(p)-N_(a)-YYY-N_(b)-ZZZ-N_(a)-n_(q)3′  (Ib);

5′n_(p)-N_(a)-XXX-N_(b)-YYY-N_(a)-n_(q)3′  (Ic); or

5′n_(p)-N_(a)-XXX-N_(b)-YYY-N_(b)-ZZZ-N_(a)-n_(q)3′  (Id).

When the sense strand is represented by formula (Ib), N_(b) representsan oligonucleotide sequence comprising 0-10, 0-7, 0-5, 0-4, 0-2, or 0modified nucleotides. Each N_(a) independently can represent anoligonucleotide sequence comprising 2-20, 2-15, or 2-10 modifiednucleotides.

When the sense strand is represented as formula (Ic), N_(b) representsan oligonucleotide sequence comprising 0-10, 0-7, 0-10, 0-7, 0-5, 0-4,0-2, or 0 modified nucleotides. Each N_(a) can independently representan oligonucleotide sequence comprising 2-20, 2-15, or 2-10 modifiednucleotides.

When the sense strand is represented as formula (Id), each N_(b)independently represents an oligonucleotide sequence comprising 0-10,0-7, 0-5, 0-4, 0-2, or 0 modified nucleotides. In one embodiment, N_(b)is 0, 1, 2, 3, 4, 5, or 6. Each N_(a) can independently represent anoligonucleotide sequence comprising 2-20, 2-15, or 2-10 modifiednucleotides.

Each of X, Y and Z may be the same or different from each other.

In other embodiments, i is 0 and j is 0, and the sense strand may berepresented by the formula:

5′n_(p)-N_(a)-YYY-N_(a)-n_(q)3′  (Ia).

When the sense strand is represented by formula (Ia), each N_(a)independently can represent an oligonucleotide sequence comprising 2-20,2-15, or 2-10 modified nucleotides.

In one embodiment, the antisense strand sequence of the RNAi may berepresented by formula (II):

5′n_(q′)-N_(a)′-(Z′Z′Z′)_(k)-N_(b)′-Y′Y′Y′-N_(b)′-(X′X′X′)_(l)-N′_(a)-n_(p)′3′  (II)

wherein:

k and l are each independently 0 or 1;

p′ and q′ are each independently 0-6;

each N_(a)′ independently represents an oligonucleotide sequencecomprising 0-25 modified nucleotides, each sequence comprising at leasttwo differently modified nucleotides;

each N_(b)′ independently represents an oligonucleotide sequencecomprising 0-10 modified nucleotides;

each n_(p)′ and n_(q)′ independently represent an overhang nucleotide;

wherein N_(b)′ and Y′ do not have the same modification; and

X′X′X′, Y′Y′Y′, and Z′Z′Z′ each independently represent one motif ofthree identical modifications on three consecutive nucleotides.

In some embodiments, the N_(a)′ or N_(b)′ comprises modifications ofalternating pattern.

The Y′Y′Y′ motif occurs at or near the cleavage site of the antisensestrand. For example, when the dsRNAi agent has a duplex region of 17-23nucleotides in length, the Y′Y′Y′ motif can occur at positions 9, 10,11; 10, 11, 12; 11, 12, 13; 12, 13, 14; or 13, 14, 15 of the antisensestrand, with the count starting from the first nucleotide, from the5′-end; or optionally, the count starting at the first paired nucleotidewithin the duplex region, from the 5′-end. In one embodiment, the Y′Y′Y′motif occurs at positions 11, 12, 13.

In certain embodiments, Y′Y′Y′ motif is all 2′-OMe modified nucleotides.

In certain embodiments, k is 1 and l is 0, or k is 0 and l is 1, or bothk and l are 1.

The antisense strand can therefore be represented by the followingformulas:

5′n_(q′)-N_(a)′-Z′Z′Z′-N_(b)′-Y′Y′Y′-N_(a)′-n_(p′)3′  (IIb);

5′n_(q′)-N_(a)′-Y′Y′Y′-N_(b)′-X′X′X′-n_(p′)3′  (IIc); or

5′n_(q′)-N_(a)′-Z′Z′Z′-N_(b)′-Y′Y′Y′-N_(b)′-X′X′X′-N_(a)′-n_(p′)3′  (IId).

When the antisense strand is represented by formula (IIb), N_(b)′represents an oligonucleotide sequence comprising 0-10, 0-7, 0-10, 0-7,0-5, 0-4, 0-2, or 0 modified nucleotides. Each N_(a)′ independentlyrepresents an oligonucleotide sequence comprising 2-20, 2-15, or 2-10modified nucleotides.

When the antisense strand is represented as formula (IIc), N_(b)′represents an oligonucleotide sequence comprising 0-10, 0-7, 0-10, 0-7,0-5, 0-4, 0-2, or 0 modified nucleotides. Each N_(a)′ independentlyrepresents an oligonucleotide sequence comprising 2-20, 2-15, or 2-10modified nucleotides.

When the antisense strand is represented as formula (IId), each N_(b)′independently represents an oligonucleotide sequence comprising 0-10,0-7, 0-10, 0-7, 0-5, 0-4, 0-2, or 0 modified nucleotides. Each N_(a)′independently represents an oligonucleotide sequence comprising 2-20,2-15, or 2-10 modified nucleotides. In one embodiment, N_(b) is 0, 1, 2,3, 4, 5, or 6.

In other embodiments, k is 0 and l is 0 and the antisense strand may berepresented by the formula:

5′n_(p′)-N_(a′)-Y′Y′Y′-N_(a′)-n_(q′)3′  (Ia).

When the antisense strand is represented as formula (IIa), each N_(a)′independently represents an oligonucleotide sequence comprising 2-20,2-15, or 2-10 modified nucleotides.

Each of X′, Y′ and Z′ may be the same or different from each other.

Each nucleotide of the sense strand and antisense strand may beindependently modified with LNA, CRN, UNA, cEt, glycol nucleic acid(GNA), hexitol nucleic acid (HNA) CeNA, 2′-methoxyethyl, 2′-O-methyl,2′-O-allyl, 2′-C-allyl, 2′-hydroxyl, or 2′-fluoro. For example, eachnucleotide of the sense strand and antisense strand is independentlymodified with 2′-O-methyl or 2′-fluoro. Each X, Y, Z, X′, Y′, and Z′, inparticular, may represent a 2′-O-methyl modification or a 2′-fluoromodification.

In some embodiments, the sense strand of the dsRNAi agent may containYYY motif occurring at 9, 10, and 11 positions of the strand when theduplex region is 21 nt, the count starting from the first nucleotidefrom the 5′-end, or optionally, the count starting at the first pairednucleotide within the duplex region, from the 5′-end; and Y represents2′-F modification. The sense strand may additionally contain XXX motifor ZZZ motifs as wing modifications at the opposite end of the duplexregion; and XXX and ZZZ each independently represents a 2′-OMemodification or 2′-F modification.

In some embodiments the antisense strand may contain Y′Y′Y′ motifoccurring at positions 11, 12, 13 of the strand, the count starting fromthe first nucleotide from the 5′-end, or optionally, the count startingat the first paired nucleotide within the duplex region, from the5′-end; and Y′ represents 2′-O-methyl modification. The antisense strandmay additionally contain X′X′X′ motif or Z′Z′Z′ motifs as wingmodifications at the opposite end of the duplex region; and X′X′X′ andZ′Z′Z′ each independently represents a 2′-OMe modification or 2′-Fmodification.

The sense strand represented by any one of the above formulas (Ia),(Ib), (Ic), and (Id) forms a duplex with an antisense strand beingrepresented by any one of formulas (IIa), (IIb), (IIc), and (IId),respectively.

Accordingly, the dsRNAi agents for use in the methods of the inventionmay comprise a sense strand and an antisense strand, each strand having14 to 30 nucleotides, the iRNA duplex represented by formula (III):

sense: 5′n_(p)-N_(a)-(XXX)_(i)-N_(b)-YYY-N_(b)-(ZZZ)-N_(a)-n_(q)3′

antisense:3′n_(p)′-N_(a)′-(X′X′X′)_(k)-N_(b)′-Y′Y′Y′-N_(b)′-(Z′Z′Z′)_(l)-N_(a)′-n_(q)′5′  (III)

wherein:

i, j, k, and l are each independently 0 or 1;

p, p′, q, and q′ are each independently 0-6;

each N_(a) and N_(a)′ independently represents an oligonucleotidesequence comprising 0-25 modified nucleotides, each sequence comprisingat least two differently modified nucleotides;

each N_(b) and N_(b)′ independently represents an oligonucleotidesequence comprising 0-10 modified nucleotides;

wherein each n_(p)′, n_(p), n_(q)′, and n_(q), each of which may or maynot be present, independently represents an overhang nucleotide; and

XXX, YYY, ZZZ, X′X′X′, Y′Y′Y′, and Z′Z′Z′ each independently representone motif of three identical modifications on three consecutivenucleotides.

In one embodiment, i is 0 and j is 0; or i is 1 and j is 0; or i is 0and j is 1; or both i and j are 0; or both i and j are 1. In anotherembodiment, k is 0 and l is 0; or k is 1 and l is 0; k is 0 and l is 1;or both k and l are 0; or both k and l are 1.

Exemplary combinations of the sense strand and antisense strand formingan iRNA duplex include the formulas below:

5′n_(p)-N_(a)-YYY-N_(a)-n_(q)3′

3′n_(p)′-N_(a)′-Y′Y′Y′-N_(a)′n_(q)′5′   (IIIa)

5′n_(p)-N_(a)-YYY-N_(b)-ZZZ-N_(a)-n_(q)3′

3′n_(p)′-N_(a)′-Y′Y′Y′-N_(b)′-Z′Z′Z′-N_(a)′n_(q)′5′   (IIIb)

5′n_(p)-N_(a)-XXX-N_(b)-YYY-N_(a)-n_(q)3′

3′n_(p)′-N_(a)′-X′X′X′-N_(b)′-Y′Y′Y′-N_(a)′-n_(q)′5′   (IIIc)

5′n_(p)-N_(a)-XXX-N_(b)-YYY-N_(b)-ZZZ-N_(a)-n_(q)3′

3′n_(p)′-N_(a)′-X′X′X′-N_(b)′-Y′Y′Y′-N_(b)′-Z′Z′Z′-N_(a)-n_(q)′5′  (IIId)

When the dsRNAi agent is represented by formula (IIIa), each N_(a)independently represents an oligonucleotide sequence comprising 2-20,2-15, or 2-10 modified nucleotides.

When the dsRNAi agent is represented by formula (IIIb), each N_(b)independently represents an oligonucleotide sequence comprising 1-10,1-7, 1-5, or 1-4 modified nucleotides. Each N_(a) independentlyrepresents an oligonucleotide sequence comprising 2-20, 2-15, or 2-10modified nucleotides.

When the dsRNAi agent is represented as formula (IIIc), each N_(b),N_(b)′ independently represents an oligonucleotide sequence comprising0-10, 0-7, 0-10, 0-7, 0-5, 0-4, 0-2, or 0 modified nucleotides. EachN_(a) independently represents an oligonucleotide sequence comprising2-20, 2-15, or 2-10 modified nucleotides.

When the dsRNAi agent is represented as formula (IIId), each N_(b),N_(b)′ independently represents an oligonucleotide sequence comprising0-10, 0-7, 0-10, 0-7, 0-5, 0-4, 0-2, or 0 modified nucleotides. EachN_(a), N_(a)′ independently represents an oligonucleotide sequencecomprising 2-20, 2-15, or 2-10 modified nucleotides. Each of N_(a),N_(a)′, N_(b), and N_(b)′ independently comprises modifications ofalternating pattern.

Each of X, Y, and Z in formulas (III), (IIIa), (IIIb), (IIIc), and(IIId) may be the same or different from each other.

When the dsRNAi agent is represented by formula (III), (IIIa), (IIIb),(IIIc), and (IIId), at least one of the Y nucleotides may form a basepair with one of the Y′ nucleotides. Alternatively, at least two of theY nucleotides form base pairs with the corresponding Y′ nucleotides; orall three of the Y nucleotides all form base pairs with thecorresponding Y′ nucleotides.

When the dsRNAi agent is represented by formula (IIIb) or (IIId), atleast one of the Z nucleotides may form a base pair with one of the Z′nucleotides. Alternatively, at least two of the Z nucleotides form basepairs with the corresponding Z′ nucleotides; or all three of the Znucleotides all form base pairs with the corresponding Z′ nucleotides.

When the dsRNAi agent is represented as formula (IIIc) or (IIId), atleast one of the X nucleotides may form a base pair with one of the X′nucleotides. Alternatively, at least two of the X nucleotides form basepairs with the corresponding X′ nucleotides; or all three of the Xnucleotides all form base pairs with the corresponding X′ nucleotides.

In certain embodiments, the modification on the Y nucleotide isdifferent than the modification on the Y′ nucleotide, the modificationon the Z nucleotide is different than the modification on the Z′nucleotide, or the modification on the X nucleotide is different thanthe modification on the X′ nucleotide.

In certain embodiments, when the dsRNAi agent is represented by formula(IIId), the N_(a) modifications are 2′-O-methyl or 2′-fluoromodifications. In other embodiments, when the RNAi agent is representedby formula (IIId), the N_(a) modifications are 2′-O-methyl or 2′-fluoromodifications and n_(p)′>0 and at least one n_(p)′ is linked to aneighboring nucleotide a via phosphorothioate linkage. In yet otherembodiments, when the RNAi agent is represented by formula (IIId), theN_(a) modifications are 2′-O-methyl or 2′-fluoro modifications, n_(p)′>0and at least one n_(p)′ is linked to a neighboring nucleotide viaphosphorothioate linkage, and the sense strand is conjugated to one ormore GalNAc derivatives attached through a bivalent or trivalentbranched linker (described below). In other embodiments, when the RNAiagent is represented by formula (IIId), the N_(a) modifications are2′-O-methyl or 2′-fluoro modifications, n_(p)′>0 and at least one n_(p)′is linked to a neighboring nucleotide via phosphorothioate linkage, thesense strand comprises at least one phosphorothioate linkage, and thesense strand is conjugated to one or more GalNAc derivatives attachedthrough a bivalent or trivalent branched linker.

In some embodiments, when the dsRNAi agent is represented by formula(IIIa), the N_(a) modifications are 2′-O-methyl or 2′-fluoromodifications, n_(p)′>0 and at least one n_(p)′ is linked to aneighboring nucleotide via phosphorothioate linkage, the sense strandcomprises at least one phosphorothioate linkage, and the sense strand isconjugated to one or more GalNAc derivatives attached through a bivalentor trivalent branched linker.

In some embodiments, the dsRNAi agent is a multimer containing at leasttwo duplexes represented by formula (III), (IIIa), (IIIb), (IIIc), and(IIId), wherein the duplexes are connected by a linker. The linker canbe cleavable or non-cleavable. Optionally, the multimer furthercomprises a ligand. Each of the duplexes can target the same gene or twodifferent genes; or each of the duplexes can target same gene at twodifferent target sites.

In some embodiments, the dsRNAi agent is a multimer containing three,four, five, six, or more duplexes represented by formula (III), (IIIa),(IIIb), (IIIc), and (IIId), wherein the duplexes are connected by alinker. The linker can be cleavable or non-cleavable. Optionally, themultimer further comprises a ligand. Each of the duplexes can target thesame gene or two different genes; or each of the duplexes can targetsame gene at two different target sites.

In one embodiment, two dsRNAi agents represented by at least one offormulas (III), (IIIa), (IIIb), (IIIc), and (IIId) are linked to eachother at the 5′ end, and one or both of the 3′ ends, and are optionallyconjugated to a ligand. Each of the agents can target the same gene ortwo different genes; or each of the agents can target same gene at twodifferent target sites.

In certain embodiments, an RNAi agent of the invention may contain a lownumber of nucleotides containing a 2′-fluoro modification, e.g., 10 orfewer nucleotides with 2′-fluoro modification. For example, the RNAiagent may contain 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 or 0 nucleotides with a2′-fluoro modification. In a specific embodiment, the RNAi agent of theinvention contains 10 nucleotides with a 2′-fluoro modification, e.g., 4nucleotides with a 2′-fluoro modification in the sense strand and 6nucleotides with a 2′-fluoro modification in the antisense strand. Inanother specific embodiment, the RNAi agent of the invention contains 6nucleotides with a 2′-fluoro modification, e.g., 4 nucleotides with a2′-fluoro modification in the sense strand and 2 nucleotides with a2′-fluoro modification in the antisense strand.

In other embodiments, an RNAi agent of the invention may contain anultra low number of nucleotides containing a 2′-fluoro modification,e.g., 2 or fewer nucleotides containing a 2′-fluoro modification. Forexample, the RNAi agent may contain 2, 1 of 0 nucleotides with a2′-fluoro modification. In a specific embodiment, the RNAi agent maycontain 2 nucleotides with a 2′-fluoro modification, e.g., 0 nucleotideswith a 2-fluoro modification in the sense strand and 2 nucleotides witha 2′-fluoro modification in the antisense strand.

Various publications describe multimeric iRNAs that can be used in themethods of the invention. Such publications include WO2007/091269, U.S.Pat. No. 7,858,769, WO2010/141511, WO2007/117686, WO2009/014887, andWO2011/031520 the entire contents of each of which are herebyincorporated herein by reference.

In certain embodiments, the compositions and methods of the disclosureinclude a vinyl phosphonate (VP) modification of an RNAi agent asdescribed herein. In exemplary embodiments, a 5′-vinyl phosphonatemodified nucleotide of the disclosure has the structure:

wherein X is O or S;

R is hydrogen, hydroxy, fluoro, or C₁₋₂₀alkoxy (e.g., methoxy orn-hexadecyloxy);

R^(5′) is ═C(H)—P(O)(OH)₂ and the double bond between the C5′ carbon andR^(5′) is in the E or Z orientation (e.g., E orientation); and

B is a nucleobase or a modified nucleobase, optionally where B isadenine, guanine, cytosine, thymine, or uracil.

In one embodiment, R^(5′) is ═C(H)—P(O)(OH)₂ and the double bond betweenthe C5′ carbon and R5′ is in the E orientation. In another embodiment, Ris methoxy and R^(5′) is ═C(H)—P(O)(OH)₂ and the double bond between theC5′ carbon and R5′ is in the E orientation. In another embodiment, X isS, R is methoxy, and R^(5′) is ═C(H)—P(O)(OH)₂ and the double bondbetween the C5′ carbon and R5′ is in the E orientation.

A vinyl phosphonate of the instant disclosure may be attached to eitherthe antisense or the sense strand of a dsRNA of the disclosure. Incertain embodiments, a on phosphonate of the instant disclosure isattached to the antisense strand of a dsRNA, optionally at the 5′ end ofthe antisense strand of the dsRNA.

Vinyl phosphonate modifications are also contemplated for thecompositions and methods of the instant disclosure. An exemplary vinylphosphonate structure includes the preceding structure, where R5′ is═C(H)—OP(O)(OH)2 and the double bond between the C5′ carbon and R5′ isin the E or Z orientation (e.g., E orientation).

As described in more detail below, the iRNA that contains conjugationsof one or more carbohydrate moieties to an iRNA can optimize one or moreproperties of the iRNA. In many cases, the carbohydrate moiety will beattached to a modified subunit of the iRNA. For example, the ribosesugar of one or more ribonucleotide subunits of an iRNA can be replacedwith another moiety, e.g., a non-carbohydrate (such as, cyclic) carrierto which is attached a carbohydrate ligand. A ribonucleotide subunit inwhich the ribose sugar of the subunit has been so replaced is referredto herein as a ribose replacement modification subunit (RRMS). A cycliccarrier may be a carbocyclic ring system, i.e., all ring atoms arecarbon atoms, or a heterocyclic ring system, i.e., one or more ringatoms may be a heteroatom, e.g., nitrogen, oxygen, sulfur. The cycliccarrier may be a monocyclic ring system, or may contain two or morerings, e.g. fused rings. The cyclic carrier may be a fully saturatedring system, or it may contain one or more double bonds.

The ligand may be attached to the polynucleotide via a carrier. Thecarriers include (i) at least one “backbone attachment point,” such as,two “backbone attachment points” and (ii) at least one “tetheringattachment point.” A “backbone attachment point” as used herein refersto a functional group, e.g. a hydroxyl group, or generally, a bondavailable for, and that is suitable for incorporation of the carrierinto the backbone, e.g., the phosphate, or modified phosphate, e.g.,sulfur containing, backbone, of a ribonucleic acid. A “tetheringattachment point” (TAP) in some embodiments refers to a constituent ringatom of the cyclic carrier, e.g., a carbon atom or a heteroatom(distinct from an atom which provides a backbone attachment point), thatconnects a selected moiety. The moiety can be, e.g., a carbohydrate,e.g. monosaccharide, disaccharide, trisaccharide, tetrasaccharide,oligosaccharide, or polysaccharide. Optionally, the selected moiety isconnected by an intervening tether to the cyclic carrier. Thus, thecyclic carrier will often include a functional group, e.g., an aminogroup, or generally, provide a bond, that is suitable for incorporationor tethering of another chemical entity, e.g., a ligand to theconstituent ring.

The iRNA may be conjugated to a ligand via a carrier, wherein thecarrier can be cyclic group or acyclic group. In some embodiments, thecyclic group is selected from pyrrolidinyl, pyrazolinyl, pyrazolidinyl,imidazolinyl, imidazolidinyl, piperidinyl, piperazinyl, [1,3]dioxolane,oxazolidinyl, isoxazolidinyl, morpholinyl, thiazolidinyl,isothiazolidinyl, quinoxalinyl, pyridazinonyl, tetrahydrofuryl, anddecalin. In some embodiments, the acyclic group is a serinol backbone ordiethanolamine backbone.

i. Thermally Destabilizing Modifications

In certain embodiments, a dsRNA molecule can be optimized for RNAinterference by incorporating thermally destabilizing modifications inthe seed region of the antisense strand. As used herein “seed region”means at positions 2-9 of the 5′-end of the referenced strand or atpositions 2-8 of the 5′-end of the referenced strand. For example,thermally destabilizing modifications can be incorporated in the seedregion of the antisense strand to reduce or inhibit off-target genesilencing.

The term “thermally destabilizing modification(s)” includesmodification(s) that would result with a dsRNA with a lower overallmelting temperature (Tm) than the Tm of the dsRNA without having suchmodification(s). For example, the thermally destabilizingmodification(s) can decrease the Tm of the dsRNA by 1-4° C., such asone, two, three or four degrees Celcius. And, the term “thermallydestabilizing nucleotide” refers to a nucleotide containing one or morethermally destabilizing modifications.

It has been discovered that dsRNAs with an antisense strand comprisingat least one thermally destabilizing modification of the duplex withinthe first 9 nucleotide positions, counting from the 5′ end, of theantisense strand have reduced off-target gene silencing activity.Accordingly, in some embodiments, the antisense strand comprises atleast one (e.g., one, two, three, four, five or more) thermallydestabilizing modification of the duplex within the first 9 nucleotidepositions of the 5′ region of the antisense strand. In some embodiments,one or more thermally destabilizing modification(s) of the duplex is/arelocated in positions 2-9, such as, positions 4-8, from the 5′-end of theantisense strand. In some further embodiments, the thermallydestabilizing modification(s) of the duplex is/are located at position6, 7 or 8 from the 5′-end of the antisense strand. In still some furtherembodiments, the thermally destabilizing modification of the duplex islocated at position 7 from the 5′-end of the antisense strand. In someembodiments, the thermally destabilizing modification of the duplex islocated at position 2, 3, 4, 5 or 9 from the 5′-end of the antisensestrand.

The thermally destabilizing modifications can include, but are notlimited to, abasic modification; mismatch with the opposing nucleotidein the opposing strand; and sugar modification such as 2′-deoxymodification, acyclic nucleotide, e.g., unlocked nucleic acids (UNA) orglycol nucleic acid (GNA); and 2′-5′-linked ribonucleotides (“3′-RNA”).

An iRNA agent comprises a sense strand and an antisense strand, eachstrand having 14 to 40 nucleotides. The RNAi agent may be represented byformula (L):

In formula (L), B1, B2, B3, B1′, B2′, B3′, and B4′ each areindependently a nucleotide containing a modification selected from thegroup consisting of 2′-O-alkyl, 2′-substituted alkoxy, 2′-substitutedalkyl, 2′-halo, ENA, and BNA/LNA. In one embodiment, B1, B2, B3, B1′,B2′, B3′, and B4′ each contain 2′-OMe modifications. In one embodiment,B1, B2, B3, B1′, B2′, B3′, and B4′ each contain 2′-OMe or 2′-Fmodifications. In one embodiment, at least one of B1, B2, B3, B1′, B2′,B3′, and B4′ contain 2′-O—N-methylacetamido (2′-O-NMA) modification.

C1 is a thermally destabilizing nucleotide placed at a site opposite tothe seed region of the antisense strand (i.e., at positions 2-8 of the5′-end of the antisense strand or at positions 2-9 of the 5′-end of thereferenced strand). For example, C1 is at a position of the sense strandthat pairs with a nucleotide at positions 2-8 of the 5′-end of theantisense strand. In one example, C1 is at position 15 from the 5′-endof the sense strand. C1 nucleotide bears the thermally destabilizingmodification which can include abasic modification; mismatch with theopposing nucleotide in the duplex; and sugar modification such as2′-deoxy modification or acyclic nucleotide e.g., unlocked nucleic acids(UNA) or glycerol nucleic acid (GNA). In one embodiment, C1 hasthermally destabilizing modification selected from the group consistingof: i) mismatch with the opposing nucleotide in the antisense strand;ii) abasic modification selected from the group consisting of:

and iii) sugar modification selected from the group consisting of:

wherein B is a modified or unmodified nucleobase, R¹ and R²independently are H, halogen, OR₃, or alkyl; and R₃ is H, alkyl,cycloalkyl, aryl, aralkyl, heteroaryl or sugar. In one embodiment, thethermally destabilizing modification in C1 is a mismatch selected fromthe group consisting of G:G, G:A, G:U, G:T, A:A, A:C, C:C, C:U, C:T,U:U, T:T, and U:T; and optionally, at least one nucleobase in themismatch pair is a 2′-deoxy nucleobase. In one example, the thermallydestabilizing modification in C1 is GNA or

T1, T1′, T2′, and T3′ each independently represent a nucleotidecomprising a modification providing the nucleotide a steric bulk that isless or equal to the steric bulk of a 2′-OMe modification. A steric bulkrefers to the sum of steric effects of a modification. Methods fordetermining steric effects of a modification of a nucleotide are knownto one skilled in the art. The modification can be at the 2′ position ofa ribose sugar of the nucleotide, or a modification to a non-ribosenucleotide, acyclic nucleotide, or the backbone of the nucleotide thatis similar or equivalent to the 2′ position of the ribose sugar, andprovides the nucleotide a steric bulk that is less than or equal to thesteric bulk of a 2′-OMe modification. For example, T1, T1′, T2′, and T3′are each independently selected from DNA, RNA, LNA, 2′-F, and2′-F-5′-methyl. In one embodiment, T1 is DNA. In one embodiment, T1′ isDNA, RNA or LNA. In one embodiment, T2′ is DNA or RNA. In oneembodiment, T3′ is DNA or RNA.n¹, n³, and q¹ are independently 4 to 15 nucleotides in length.n⁵, q³, and q⁷ are independently 1-6 nucleotide(s) in length.n⁴, q², and q⁶ are independently 1-3 nucleotide(s) in length;alternatively, n⁴ is 0.q⁵ is independently 0-10 nucleotide(s) in length.n² and q⁴ are independently 0-3 nucleotide(s) in length.

Alternatively, n⁴ is 0-3 nucleotide(s) in length.

In one embodiment, n⁴ can be 0. In one example, n⁴ is 0, and q² and q⁶are 1. In another example, n⁴ is 0, and q² and q⁶ are 1, with twophosphorothioate internucleotide linkage modifications within position1-5 of the sense strand (counting from the 5′-end of the sense strand),and two phosphorothioate internucleotide linkage modifications atpositions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end of the antisense strand).

In one embodiment, n⁴, q², and q⁶ are each 1.

In one embodiment, n², n⁴, q², q⁴, and q are each 1.

In one embodiment, C1 is at position 14-17 of the 5′-end of the sensestrand, when the sense strand is 19-22 nucleotides in length, and n⁴is 1. In one embodiment, C1 is at position 15 of the 5′-end of the sensestrand

In one embodiment, T3′ starts at position 2 from the 5′ end of theantisense strand. In one example, T3′ is at position 2 from the 5′ endof the antisense strand and q⁶ is equal to 1.

In one embodiment, T1′ starts at position 14 from the 5′ end of theantisense strand. In one example, T1′ is at position 14 from the 5′ endof the antisense strand and q² is equal to 1.

In an exemplary embodiment, T3′ starts from position 2 from the 5′ endof the antisense strand and T1′ starts from position 14 from the 5′ endof the antisense strand. In one example, T3′ starts from position 2 fromthe 5′ end of the antisense strand and q⁶ is equal to 1 and T1′ startsfrom position 14 from the 5′ end of the antisense strand and q² is equalto 1.

In one embodiment, T1′ and T3′ are separated by 11 nucleotides in length(i.e. not counting the T1′ and T3′ nucleotides).

In one embodiment, T1′ is at position 14 from the 5′ end of theantisense strand. In one example, T1′ is at position 14 from the 5′ endof the antisense strand and q² is equal to 1, and the modification atthe 2′ position or positions in a non-ribose, acyclic or backbone thatprovide less steric bulk than a 2′-OMe ribose.

In one embodiment, T3′ is at position 2 from the 5′ end of the antisensestrand. In one example, T3′ is at position 2 from the 5′ end of theantisense strand and q⁶ is equal to 1, and the modification at the 2′position or positions in a non-ribose, acyclic or backbone that provideless than or equal to steric bulk than a 2′-OMe ribose.

In one embodiment, T1 is at the cleavage site of the sense strand. Inone example, T1 is at position 11 from the 5′ end of the sense strand,when the sense strand is 19-22 nucleotides in length, and n² is 1. In anexemplary embodiment, T1 is at the cleavage site of the sense strand atposition 11 from the 5′ end of the sense strand, when the sense strandis 19-22 nucleotides in length, and n² is 1,

In one embodiment, T2′ starts at position 6 from the 5′ end of theantisense strand. In one example, T2′ is at positions 6-10 from the 5′end of the antisense strand, and q⁴ is 1.

In an exemplary embodiment, T1 is at the cleavage site of the sensestrand, for instance, at position 11 from the 5′ end of the sensestrand, when the sense strand is 19-22 nucleotides in length, and n² is1; T1′ is at position 14 from the 5′ end of the antisense strand, and q²is equal to 1, and the modification to T1′ is at the 2′ position of aribose sugar or at positions in a non-ribose, acyclic or backbone thatprovide less steric bulk than a 2′-OMe ribose; T2′ is at positions 6-10from the 5′ end of the antisense strand, and q⁴ is 1; and T3′ is atposition 2 from the 5′ end of the antisense strand, and q⁶ is equal to1, and the modification to T3′ is at the 2′ position or at positions ina non-ribose, acyclic or backbone that provide less than or equal tosteric bulk than a 2′-OMe ribose.

In one embodiment, T2′ starts at position 8 from the 5′ end of theantisense strand. In one example, T2′ starts at position 8 from the 5′end of the antisense strand, and q⁴ is 2.

In one embodiment, T2′ starts at position 9 from the 5′ end of theantisense strand. In one example, T2′ is at position 9 from the 5′ endof the antisense strand, and q⁴ is 1.

In one embodiment, B1′ is 2′-OMe or 2′-F, q¹ is 9, T1′ is 2′-F, q² is 1,B2′ is 2′-OMe or 2′-F, q³ is 4, T2′ is 2′-F, q⁴ is 1, B3′ is 2′-OMe or2′-F, q⁵ is 6, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-OMe, and q⁷ is 1; withtwo phosphorothioate internucleotide linkage modifications withinpositions 1-5 of the sense strand (counting from the 5′-end of the sensestrand), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end of the antisense strand).

In one embodiment, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 1, B3′ is 2′-OMe or 2′-F, q⁵ is 6, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within positions 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within positions 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 6, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 7, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 6, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 7, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within positions 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 1, B3′ is 2′-OMe or 2′-F, q⁵ is 6, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 1, B3′ is 2′-OMe or 2′-F, q⁵ is 6, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within positions 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 5, T2′is 2′-F, q⁴ is 1, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; optionally with at least 2 additional TT atthe 3′-end of the antisense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 5, T2′is 2′-F, q⁴ is 1, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; optionally with at least 2 additional TT atthe 3′-end of the antisense strand; with two phosphorothioateinternucleotide linkage modifications within positions 1-5 of the sensestrand (counting from the 5′-end of the sense strand), and twophosphorothioate internucleotide linkage modifications at positions 1and 2 and two phosphorothioate internucleotide linkage modificationswithin positions 18-23 of the antisense strand (counting from the 5′-endof the antisense strand).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within positions 1-5 of the sense strand (counting fromthe 5′-end), and two phosphorothioate internucleotide linkagemodifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1. In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is8, T1 is 2′F, n² is 3, B2 is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵is 3, B1′ is 2′-OMe or 2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is2′-OMe or 2′-F, q³ is 4, T2′ is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵is 5, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F, and q⁷ is 1; with twophosphorothioate internucleotide linkage modifications within positions1-5 of the sense strand (counting from the 5′-end of the sense strand),and two phosphorothioate internucleotide linkage modifications atpositions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end of the antisense strand).

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within positions 1-5 of the sense strand (counting fromthe 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand).

The RNAi agent can comprise a phosphorus-containing group at the 5′-endof the sense strand or antisense strand. The 5′-endphosphorus-containing group can be 5′-end phosphate (5′-P), 5′-endphosphorothioate (5′-PS), 5′-end phosphorodithioate (5′-PS₂), 5′-endvinylphosphonate (5′-VP), 5′-end methylphosphonate (MePhos), or5′-deoxy-5′-C-malonyl

When the 5′-end phosphorus-containing group is 5′-end vinylphosphonate(5′-VP), the 5′-VP can be either 5′-E-VP isomer (i.e.,trans-vinylphosphonate,

5′-Z-VP isomer (i.e., cis-vinylphosphonate,

or mixtures thereof.In one embodiment, the RNAi agent comprises a phosphorus-containinggroup at the 5′-end of the sense strand. In one embodiment, the RNAiagent comprises a phosphorus-containing group at the 5′-end of theantisense strand.

In one embodiment, the RNAi agent comprises a 5′-P. In one embodiment,the RNAi agent comprises a 5′-P in the antisense strand.

In one embodiment, the RNAi agent comprises a 5′-PS. In one embodiment,the RNAi agent comprises a 5′-PS in the antisense strand.

In one embodiment, the RNAi agent comprises a 5′-VP. In one embodiment,the RNAi agent comprises a 5′-VP in the antisense strand. In oneembodiment, the RNAi agent comprises a 5′-E-VP in the antisense strand.In one embodiment, the RNAi agent comprises a 5′-Z-VP in the antisensestrand.

In one embodiment, the RNAi agent comprises a 5′-PS₂. In one embodiment,the RNAi agent comprises a 5′-PS₂ in the antisense strand.

In one embodiment, the RNAi agent comprises a 5′-PS₂. In one embodiment,the RNAi agent comprises a 5′-deoxy-5′-C-malonyl in the antisensestrand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-VP. The5′-VP may be 5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁵ is 1,B4′ is 2′-OMe, and q⁷ is 1. The RNAi agent also comprises a5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-VP. The 5′-VP may be5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁵ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1. The dsRNA agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-VP. The 5′-VPmay be 5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1. The RNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1. The RNAi agent also comprises a5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, 10 q³ is 4,q⁴ is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-VP. The 5′-VP maybe 5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1. The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1. The RNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1. The RNAi agent also comprises a 5′-VP. The5′-VP may be 5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1. The dsRNAi RNA agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1. The RNAi agent also comprises a5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n^(d) is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-VP. The 5′-VP may be5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1. The RNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1. The RNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1. The RNAi agent also comprises a 5′-VP. The 5′-VP may be5′-E-VP, 5′-Z-VP, or combination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1. The RNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1. The RNAi agent also comprises a 5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-P.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-PS.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-VP. The 5′-VP may be 5′-E-VP, 5′-Z-VP, orcombination thereof.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-PS₂.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-deoxy-5′-C-malonyl.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-P and a targeting ligand. Inone embodiment, the 5′-P is at the 5′-end of the antisense strand, andthe targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS and a targeting ligand.In one embodiment, the 5′-PS is at the 5′-end of the antisense strand,and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-VP (e.g., a 5′-E-VP,5′-Z-VP, or combination thereof), and a targeting ligand.

In one embodiment, the 5′-VP is at the 5′-end of the antisense strand,and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS₂ and a targeting ligand.In one embodiment, the 5′-PS₂ is at the 5′-end of the antisense strand,and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-deoxy-5′-C-malonyl and atargeting ligand. In one embodiment, the 5′-deoxy-5′-C-malonyl is at the5′-end of the antisense strand, and the targeting ligand is at the3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-P and a targetingligand. In one embodiment, the 5′-P is at the 5′-end of the antisensestrand, and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-PS and a targetingligand. In one embodiment, the 5′-PS is at the 5′-end of the antisensestrand, and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-VP (e.g., a5′-E-VP, 5′-Z-VP, or combination thereof) and a targeting ligand. In oneembodiment, the 5′-VP is at the 5′-end of the antisense strand, and thetargeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-PS₂ and a targetingligand. In one embodiment, the 5′-PS₂ is at the 5′-end of the antisensestrand, and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is2′-OMe, and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end), and two phosphorothioate internucleotide linkage modificationsat positions 1 and 2 and two phosphorothioate internucleotide linkagemodifications within positions 18-23 of the antisense strand (countingfrom the 5′-end). The RNAi agent also comprises a 5′-deoxy-5′-C-malonyland a targeting ligand. In one embodiment, the 5′-deoxy-5′-C-malonyl isat the 5′-end of the antisense strand, and the targeting ligand is atthe 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-P and a targeting ligand. Inone embodiment, the 5′-P is at the 5′-end of the antisense strand, andthe targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS and a targeting ligand.In one embodiment, the 5′-PS is at the 5′-end of the antisense strand,and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-VP (e.g., a 5′-E-VP,5′-Z-VP, or combination thereof) and a targeting ligand. In oneembodiment, the 5′-VP is at the 5′-end of the antisense strand, and thetargeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-PS₂ and a targeting ligand.In one embodiment, the 5′-PS₂ is at the 5′-end of the antisense strand,and the targeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, T2′is 2′-F, q⁴ is 2, B3′ is 2′-OMe or 2′-F, q⁵ is 5, T3′ is 2′-F, q⁶ is 1,B4′ is 2′-F, and q⁷ is 1; with two phosphorothioate internucleotidelinkage modifications within position 1-5 of the sense strand (countingfrom the 5′-end of the sense strand), and two phosphorothioateinternucleotide linkage modifications at positions 1 and 2 and twophosphorothioate internucleotide linkage modifications within positions18-23 of the antisense strand (counting from the 5′-end of the antisensestrand). The RNAi agent also comprises a 5′-deoxy-5′-C-malonyl and atargeting ligand. In one embodiment, the 5′-deoxy-5′-C-malonyl is at the5′-end of the antisense strand, and the targeting ligand is at the3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-P and a targeting ligand. In oneembodiment, the 5′-P is at the 5′-end of the antisense strand, and thetargeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-PS and a targeting ligand. In oneembodiment, the 5′-PS is at the 5′-end of the antisense strand, and thetargeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-VP (e.g., a 5′-E-VP, 5′-Z-VP, orcombination thereof) and a targeting ligand. In one embodiment, the5′-VP is at the 5′-end of the antisense strand, and the targeting ligandis at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-PS₂ and a targeting ligand. In oneembodiment, the 5′-PS₂ is at the 5′-end of the antisense strand, and thetargeting ligand is at the 3′-end of the sense strand.

In one embodiment, B1 is 2′-OMe or 2′-F, n¹ is 8, T1 is 2′F, n² is 3, B2is 2′-OMe, n³ is 7, n⁴ is 0, B3 is 2′-OMe, n⁵ is 3, B1′ is 2′-OMe or2′-F, q¹ is 9, T1′ is 2′-F, q² is 1, B2′ is 2′-OMe or 2′-F, q³ is 4, q⁴is 0, B3′ is 2′-OMe or 2′-F, q⁵ is 7, T3′ is 2′-F, q⁶ is 1, B4′ is 2′-F,and q⁷ is 1; with two phosphorothioate internucleotide linkagemodifications within position 1-5 of the sense strand (counting from the5′-end of the sense strand), and two phosphorothioate internucleotidelinkage modifications at positions 1 and 2 and two phosphorothioateinternucleotide linkage modifications within positions 18-23 of theantisense strand (counting from the 5′-end of the antisense strand). TheRNAi agent also comprises a 5′-deoxy-5′-C-malonyl and a targetingligand. In one embodiment, the 5′-deoxy-5′-C-malonyl is at the 5′-end ofthe antisense strand, and the targeting ligand is at the 3′-end of thesense strand.

In a particular embodiment, an RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;        -   (ii) an ASGPR ligand attached to the 3′-end, wherein said            ASGPR ligand comprises three GalNAc derivatives attached            through a trivalent branched linker; and        -   (iii) 2′-F modifications at positions 1, 3, 5, 7, 9 to 11,            13, 17, 19, and 21, and 2′-OMe modifications at positions 2,            4, 6, 8, 12, 14 to 16, 18, and 20 (counting from the 5′            end); and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;        -   (ii) 2′-OMe modifications at positions 1, 3, 5, 9, 11 to 13,            15, 17, 19, 21, and 23, and 2′F modifications at positions            2, 4, 6 to 8, 10, 14, 16, 18, 20, and 22 (counting from the            5′ end); and        -   (iii) phosphorothioate internucleotide linkages between            nucleotide positions 21 and 22, and between nucleotide            positions 22 and 23 (counting from the 5′ end);    -   wherein the dsRNA agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, an RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:    -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-F modifications at positions 1, 3, 5, 7, 9 to 11,            13, 15, 17, 19, and 21, and 2′-OMe modifications at            positions 2, 4, 6, 8, 12, 14, 16, 18, and 20 (counting from            the 5′ end); and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3, 5, 7, 9, 11 to 13,        15, 17, 19, and 21 to 23, and 2′F modifications at positions 2,        4, 6, 8, 10, 14, 16, 18, and 20 (counting from the 5′ end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1 to 6, 8, 10, and            12 to 21, 2′-F modifications at positions 7, and 9, and a            deoxy-nucleotide (e.g. dT) at position 11 (counting from the            5′ end); and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3, 7, 9, 11, 13, 15,        17, and 19 to 23, and 2′-F modifications at positions 2, 4 to 6,        8, 10, 12, 14, 16, and 18 (counting from the 5′ end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1 to 6, 8, 10, 12,            14, and 16 to 21, and 2′-F modifications at positions 7, 9,            11, 13, and 15; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 5, 7, 9, 11, 13, 15,        17, 19, and 21 to 23, and 2′-F modifications at positions 2 to        4, 6, 8, 10, 12, 14, 16, 18, and 20 (counting from the 5′ end);        and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1 to 9, and 12 to            21, and 2′-F modifications at positions 10, and 11; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3, 5, 7, 9, 11 to 13,        15, 17, 19, and 21 to 23, and 2′-F modifications at positions 2,        4, 6, 8, 10, 14, 16, 18, and 20 (counting from the 5′ end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-F modifications at positions 1, 3, 5, 7, 9 to 11,            and 13, and 2′-OMe modifications at positions 2, 4, 6, 8,            12, and 14 to 21; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3, 5 to 7, 9, 11 to        13, 15, 17 to 19, and 21 to 23, and 2′-F modifications at        positions 2, 4, 8, 10, 14, 16, and 20 (counting from the 5′        end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1, 2, 4, 6, 8, 12,            14, 15, 17, and 19 to 21, and 2′-F modifications at            positions 3, 5, 7, 9 to 11, 13, 16, and 18; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 25 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 4, 6, 7, 9, 11 to 13,        15, 17, and 19 to 23, 2′-F modifications at positions 2, 3, 5,        8, 10, 14, 16, and 18, and desoxy-nucleotides (e.g. dT) at        positions 24 and 25 (counting from the 5′ end); and        -   (iii) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, between nucleotide positions 2            and 3, between nucleotide positions 21 and 22, and between            nucleotide positions 22 and 23 (counting from the 5′ end);            wherein the RNAi agents have a four nucleotide overhang at            the 3′-end of the antisense strand, and a blunt end at the            5′-end of the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1 to 6, 8, and 12 to            21, and 2′-F modifications at positions 7, and 9 to 11; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3 to 5, 7, 8, 10 to        13, 15, and 17 to 23, and 2′-F modifications at positions 2, 6,        9, 14, and 16 (counting from the 5′ end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1 to 6, 8, and 12 to            21, and 2′-F modifications at positions 7, and 9 to 11; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 23 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3 to 5, 7, 10 to 13,        15, and 17 to 23, and 2′-F modifications at positions 2, 6, 8,        9, 14, and 16 (counting from the 5′ end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 21 and 22, and between        nucleotide positions 22 and 23 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In another particular embodiment, a RNAi agent of the present inventioncomprises:

-   -   (a) a sense strand having:        -   (i) a length of 19 nucleotides;    -   (ii) an ASGPR ligand attached to the 3′-end, wherein said ASGPR        ligand comprises three GalNAc derivatives attached through a        trivalent branched linker;        -   (iii) 2′-OMe modifications at positions 1 to 4, 6, and 10 to            19, and 2′-F modifications at positions 5, and 7 to 9; and        -   (iv) phosphorothioate internucleotide linkages between            nucleotide positions 1 and 2, and between nucleotide            positions 2 and 3 (counting from the 5′ end);    -   and    -   (b) an antisense strand having:        -   (i) a length of 21 nucleotides;    -   (ii) 2′-OMe modifications at positions 1, 3 to 5, 7, 10 to 13,        15, and 17 to 21, and 2′-F modifications at positions 2, 6, 8,        9, 14, and 16 (counting from the 5′ end); and    -   (iii) phosphorothioate internucleotide linkages between        nucleotide positions 1 and 2, between nucleotide positions 2 and        3, between nucleotide positions 19 and 20, and between        nucleotide positions 20 and 21 (counting from the 5′ end);        wherein the RNAi agents have a two nucleotide overhang at the        3′-end of the antisense strand, and a blunt end at the 5′-end of        the antisense strand.

In certain embodiments, the iRNA for use in the methods of the inventionis an agent selected from agents listed in any one of Tables 2-3. Theseagents may further comprise a ligand.

III. iRNAs Conjugated to Ligands

Another modification of the RNA of an iRNA of the invention involveschemically linking to the iRNA one or more ligands, moieties orconjugates that enhance the activity, cellular distribution, or cellularuptake of the iRNA e.g., into a cell. Such moieties include but are notlimited to lipid moieties such as a cholesterol moiety (Letsinger etal., Proc. Natl. Acid. Sci. USA, 1989, 86: 6553-6556). In otherembodiments, the ligand is cholic acid (Manoharan et al., Biorg. Med.Chem. Let., 1994, 4:1053-1060), a thioether, e.g., beryl-S-tritylthiol(Manoharan et al., Ann. N.Y. Acad. Sci., 1992, 660:306-309; Manoharan etal., Biorg. Med. Chem. Let., 1993, 3:2765-2770), a thiocholesterol(Oberhauser et al., Nucl. Acids Res., 1992, 20:533-538), an aliphaticchain, e.g., dodecandiol or undecyl residues (Saison-Behmoaras et al.,EMBO J, 1991, 10:1111-1118; Kabanov et al., FEBS Lett., 1990,259:327-330; Svinarchuk et al., Biochimie, 1993, 75:49-54), aphospholipid, e.g., di-hexadecyl-rac-glycerol or triethyl-ammonium1,2-di-O-hexadecyl-rac-glycero-3-phosphonate (Manoharan et al.,Tetrahedron Lett., 1995, 36:3651-3654; Shea et al., Nucl. Acids Res.,1990, 18:3777-3783), a polyamine or a polyethylene glycol chain(Manoharan et al., Nucleosides & Nucleotides, 1995, 14:969-973), oradamantane acetic acid (Manoharan et al., Tetrahedron Lett., 1995,36:3651-3654), a palmityl moiety (Mishra et al., Biochim. Biophys. Acta,1995, 1264:229-237), or an octadecylamine orhexylamino-carbonyloxycholesterol moiety (Crooke et al., J. Pharmacol.Exp. Ther., 1996, 277:923-937).

In certain embodiments, a ligand alters the distribution, targeting, orlifetime of an iRNA agent into which it is incorporated. In certainembodiments a ligand provides an enhanced affinity for a selectedtarget, e.g., molecule, cell or cell type, compartment, e.g., a cellularor organ compartment, tissue, organ or region of the body, as, e.g.,compared to a species absent such a ligand. In some embodiments, ligandsdo not take part in duplex pairing in a duplexed nucleic acid.

Ligands can include a naturally occurring substance, such as a protein(e.g., human serum albumin (HSA), low-density lipoprotein (LDL), orglobulin); carbohydrate (e.g., a dextran, pullulan, chitin, chitosan,inulin, cyclodextrin, N-acetylglucosamine, N-acetylgalactosamine, orhyaluronic acid); or a lipid. The ligand can also be a recombinant orsynthetic molecule, such as a synthetic polymer, e.g., a syntheticpolyamino acid. Examples of polyamino acids include polyamino acid is apolylysine (PLL), poly L-aspartic acid, poly L-glutamic acid,styrene-maleic acid anhydride copolymer, poly(L-lactide-co-glycolide)copolymer, divinyl ether-maleic anhydride copolymer,N-(2-hydroxypropyl)methacrylamide copolymer (HMPA), polyethylene glycol(PEG), polyvinyl alcohol (PVA), polyurethane, poly(2-ethylacrylic acid),N-isopropylacrylamide polymers, or polyphosphazene. Example ofpolyamines include: polyethylenimine, polylysine (PLL), spermine,spermidine, polyamine, pseudopeptide-polyamine, peptidomimeticpolyamine, dendrimer polyamine, arginine, amidine, protamine, cationiclipid, cationic porphyrin, quaternary salt of a polyamine, or an alphahelical peptide.

Ligands can also include targeting groups, e.g., a cell or tissuetargeting agent, e.g., a lectin, glycoprotein, lipid or protein, e.g.,an antibody, that binds to a specified cell type such as a kidney cell.A targeting group can be a thyrotropin, melanotropin, lectin,glycoprotein, surfactant protein A, Mucin carbohydrate, multivalentlactose, multivalent galactose, N-acetyl-galactosamine,N-acetyl-glucosamine multivalent mannose, multivalent fucose,glycosylated polyaminoacids, multivalent galactose, transferrin,bisphosphonate, polyglutamate, polyaspartate, a lipid, cholesterol, asteroid, bile acid, folate, vitamin B12, vitamin A, biotin, or an RGDpeptide or RGD peptide mimetic. In certain embodiments, the ligand is amultivalent galactose, e.g., an N-acetyl-galactosamine.

Other examples of ligands include dyes, intercalating agents (e.g.acridines), cross-linkers (e.g. psoralene, mitomycin C), porphyrins(TPPC4, texaphyrin, Sapphyrin), polycyclic aromatic hydrocarbons (e.g.,phenazine, dihydrophenazine), artificial endonucleases (e.g. EDTA),lipophilic molecules, e.g., cholesterol, cholic acid, adamantane aceticacid, 1-pyrene butyric acid, dihydrotestosterone,1,3-Bis-O(hexadecyl)glycerol, geranyloxyhexyl group, hexadecylglycerol,borneol, menthol, 1,3-propanediol, heptadecyl group, palmitic acid,myristic acid, O3-(oleoyl)lithocholic acid, O3-(oleoyl)cholenic acid,dimethoxytrityl, or phenoxazine) and peptide conjugates (e.g.,antennapedia peptide, Tat peptide), alkylating agents, phosphate, amino,mercapto, PEG (e.g., PEG-40K), MPEG, [MPEG]₂, polyamino, alkyl,substituted alkyl, radiolabeled markers, enzymes, haptens (e.g. biotin),transport/absorption facilitators (e.g., aspirin, vitamin E, folicacid), synthetic ribonucleases (e.g., imidazole, bisimidazole,histamine, imidazole clusters, acridine-imidazole conjugates, Eu3+complexes of tetraazamacrocycles), dinitrophenyl, HRP, or AP.

Ligands can be proteins, e.g., glycoproteins, or peptides, e.g.,molecules having a specific affinity for a co-ligand, or antibodiese.g., an antibody, that binds to a specified cell type such as a hepaticcell. Ligands can also include hormones and hormone receptors. They canalso include non-peptidic species, such as lipids, lectins,carbohydrates, vitamins, cofactors, multivalent lactose, multivalentgalactose, N-acetyl-galactosamine, N-acetyl-glucosamine multivalentmannose, or multivalent fucose. The ligand can be, for example, alipopolysaccharide, an activator of p38 MAP kinase, or an activator ofNF-κB.

The ligand can be a substance, e.g., a drug, which can increase theuptake of the iRNA agent into the cell, for example, by disrupting thecell's cytoskeleton, e.g., by disrupting the cell's microtubules,microfilaments, or intermediate filaments. The drug can be, for example,taxol, vincristine, vinblastine, cytochalasin, nocodazole,japlakinolide, latrunculin A, phalloidin, swinholide A, indanocine, ormyoservin.

In some embodiments, a ligand attached to an iRNA as described hereinacts as a pharmacokinetic modulator (PK modulator). PK modulatorsinclude lipophiles, bile acids, steroids, phospholipid analogues,peptides, protein binding agents, PEG, vitamins, etc. Exemplary PKmodulators include, but are not limited to, cholesterol, fatty acids,cholic acid, lithocholic acid, dialkylglycerides, diacylglyceride,phospholipids, sphingolipids, naproxen, ibuprofen, vitamin E, biotin.Oligonucleotides that comprise a number of phosphorothioate linkages arealso known to bind to serum protein, thus short oligonucleotides, e.g.,oligonucleotides of about 5 bases, 10 bases, 15 bases, or 20 bases,comprising multiple of phosphorothioate linkages in the backbone arealso amenable to the present invention as ligands (e.g. as PK modulatingligands). In addition, aptamers that bind serum components (e.g. serumproteins) are also suitable for use as PK modulating ligands in theembodiments described herein.

Ligand-conjugated iRNAs of the invention may be synthesized by the useof an oligonucleotide that bears a pendant reactive functionality, suchas that derived from the attachment of a linking molecule onto theoligonucleotide (described below). This reactive oligonucleotide may bereacted directly with commercially-available ligands, ligands that aresynthesized bearing any of a variety of protecting groups, or ligandsthat have a linking moiety attached thereto.

The oligonucleotides used in the conjugates of the present invention maybe conveniently and routinely made through the well-known technique ofsolid-phase synthesis. Equipment for such synthesis is sold by severalvendors including, for example, Applied Biosystems® (Foster City,Calif.). Any other methods for such synthesis known in the art mayadditionally or alternatively be employed. It is also known to usesimilar techniques to prepare other oligonucleotides, such as thephosphorothioates and alkylated derivatives.

In the ligand-conjugated iRNAs and ligand-molecule bearingsequence-specific linked nucleosides of the present invention, theoligonucleotides and oligonucleosides may be assembled on a suitable DNAsynthesizer utilizing standard nucleotide or nucleoside precursors, ornucleotide or nucleoside conjugate precursors that already bear thelinking moiety, ligand-nucleotide or nucleoside-conjugate precursorsthat already bear the ligand molecule, or non-nucleoside ligand-bearingbuilding blocks.

When using nucleotide-conjugate precursors that already bear a linkingmoiety, the synthesis of the sequence-specific linked nucleosides istypically completed, and the ligand molecule is then reacted with thelinking moiety to form the ligand-conjugated oligonucleotide. In someembodiments, the oligonucleotides or linked nucleosides of the presentinvention are synthesized by an automated synthesizer usingphosphoramidites derived from ligand-nucleoside conjugates in additionto the standard phosphoramidites and non-standard phosphoramidites thatare commercially available and routinely used in oligonucleotidesynthesis.

A. Lipid Conjugates

In certain embodiments, the ligand or conjugate is a lipid orlipid-based molecule. Such a lipid or lipid-based molecule may bind aserum protein, e.g., human serum albumin (HSA). An HSA binding ligandallows for distribution of the conjugate to a target tissue, e.g., anon-kidney target tissue of the body. For example, the target tissue canbe the liver, including parenchymal cells of the liver. Other moleculesthat can bind HSA can also be used as ligands. For example, naproxen oraspirin can be used. A lipid or lipid-based ligand can (a) increaseresistance to degradation of the conjugate, (b) increase targeting ortransport into a target cell or cell membrane, or (c) can be used toadjust binding to a serum protein, e.g., HSA.

A lipid based ligand can be used to inhibit, e.g., control the bindingof the conjugate to a target tissue. For example, a lipid or lipid-basedligand that binds to HSA more strongly will be less likely to betargeted to the kidney and therefore less likely to be cleared from thebody. A lipid or lipid-based ligand that binds to HSA less strongly canbe used to target the conjugate to the kidney.

In certain embodiments, the lipid based ligand binds HSA. In oneembodiment, it binds HSA with a sufficient affinity such that theconjugate will be distributed to a non-kidney tissue. However, it ispreferred that the affinity not be so strong that the HSA-ligand bindingcannot be reversed.

In other embodiments, the lipid based ligand binds HSA weakly or not atall, such that the conjugate will be distributed to the kidney. Othermoieties that target to kidney cells can also be used in place of, or inaddition to, the lipid based ligand.

In another aspect, the ligand is a moiety, e.g., a vitamin, which istaken up by a target cell, e.g., a proliferating cell. These areparticularly useful for treating disorders characterized by unwantedcell proliferation, e.g., of the malignant or non-malignant type, e.g.,cancer cells. Exemplary vitamins include vitamin A, E, and K. Otherexemplary vitamins include are B vitamin, e.g., folic acid, B12,riboflavin, biotin, pyridoxal or other vitamins or nutrients taken up bytarget cells such as liver cells. Also included are HSA and low densitylipoprotein (LDL).

B. Cell Permeation Agents

In another aspect, the ligand is a cell-permeation agent, such as, ahelical cell-permeation agent. In one embodiment, the agent isamphipathic. An exemplary agent is a peptide such as tat orantennapedia. If the agent is a peptide, it can be modified, including apeptidylmimetic, invertomers, non-peptide or pseudo-peptide linkages,and use of D-amino acids. In one embodiment, the helical agent is analpha-helical agent, for example, having a lipophilic and a lipophobicphase.

The ligand can be a peptide or peptidomimetic. A peptidomimetic (alsoreferred to herein as an oligopeptidomimetic) is a molecule capable offolding into a defined three-dimensional structure similar to a naturalpeptide. The attachment of peptide and peptidomimetics to iRNA agentscan affect pharmacokinetic distribution of the iRNA, such as byenhancing cellular recognition and absorption. The peptide orpeptidomimetic moiety can be about 5-50 amino acids long, e.g., about 5,10, 15, 20, 25, 30, 35, 40, 45, or 50 amino acids long.

A peptide or peptidomimetic can be, for example, a cell permeationpeptide, cationic peptide, amphipathic peptide, or hydrophobic peptide(e.g., consisting primarily of Tyr, Trp, or Phe). The peptide moiety canbe a dendrimer peptide, constrained peptide or crosslinked peptide. Inanother alternative, the peptide moiety can include a hydrophobicmembrane translocation sequence (MTS). An exemplary hydrophobicMTS-containing peptide is RFGF having the amino acid sequenceAAVALLPAVLLALLAP (SEQ ID NO: 15). An RFGF analogue (e.g., amino acidsequence AALLPVLLAAP (SEQ ID NO:16) containing a hydrophobic MTS canalso be a targeting moiety. The peptide moiety can be a “delivery”peptide, which can carry large polar molecules including peptides,oligonucleotides, and protein across cell membranes. For example,sequences from the HIV Tat protein (GRKKRRQRRRPPQ (SEQ ID NO: 17) andthe Drosophila Antennapedia protein (RQIKIWFQNRRMKWKK (SEQ ID NO: 18)have been found to be capable of functioning as delivery peptides. Apeptide or peptidomimetic can be encoded by a random sequence of DNA,such as a peptide identified from a phage-display library, orone-bead-one-compound (OBOC) combinatorial library (Lam et al., Nature,354:82-84, 1991). Examples of a peptide or peptidomimetic tethered to adsRNA agent via an incorporated monomer unit for cell targeting purposesis an arginine-glycine-aspartic acid (RGD)-peptide, or RGD mimic. Apeptide moiety can range in length from about 5 amino acids to about 40amino acids. The peptide moieties can have a structural modification,such as to increase stability or direct conformational properties. Anyof the structural modifications described below can be utilized.

An RGD peptide for use in the compositions and methods of the inventionmay be linear or cyclic, and may be modified, e.g., glycosylated ormethylated, to facilitate targeting to a specific tissue(s).RGD-containing peptides and peptidomimetics may include D-amino acids,as well as synthetic RGD mimics. In addition to RGD, one can use othermoieties that target the integrin ligand, such as, PECAM-1 or VEGF.

A “cell permeation peptide” is capable of permeating a cell, e.g., amicrobial cell, such as a bacterial or fungal cell, or a mammalian cell,such as a human cell. A microbial cell-permeating peptide can be, forexample, an α-helical linear peptide (e.g., LL-37 or Ceropin P1), adisulfide bond-containing peptide (e.g., α-defensin, β-defensin orbactenecin), or a peptide containing only one or two dominating aminoacids (e.g., PR-39 or indolicidin). A cell permeation peptide can alsoinclude a nuclear localization signal (NLS). For example, a cellpermeation peptide can be a bipartite amphipathic peptide, such as MPG,which is derived from the fusion peptide domain of HIV-1 gp41 and theNLS of SV40 large T antigen (Simeoni et al., Nucl. Acids Res.31:2717-2724, 2003).

C. Carbohydrate Conjugates

In some embodiments of the compositions and methods of the invention, aniRNA further comprises a carbohydrate. The carbohydrate conjugated iRNAis advantageous for the in vivo delivery of nucleic acids, as well ascompositions suitable for in vivo therapeutic use, as described herein.As used herein, “carbohydrate” refers to a compound which is either acarbohydrate per se made up of one or more monosaccharide units havingat least 6 carbon atoms (which can be linear, branched or cyclic) withan oxygen, nitrogen or sulfur atom bonded to each carbon atom; or acompound having as a part thereof a carbohydrate moiety made up of oneor more monosaccharide units each having at least six carbon atoms(which can be linear, branched or cyclic), with an oxygen, nitrogen orsulfur atom bonded to each carbon atom. Representative carbohydratesinclude the sugars (mono-, di-, tri-, and oligosaccharides containingfrom about 4, 5, 6, 7, 8, or 9 monosaccharide units), andpolysaccharides such as starches, glycogen, cellulose and polysaccharidegums. Specific monosaccharides include C5 and above (e.g., C5, C6, C7,or C8) sugars; di- and trisaccharides include sugars having two or threemonosaccharide units (e.g., C5, C6, C7, or C8).

In certain embodiments, a carbohydrate conjugate for use in thecompositions and methods of the invention is a monosaccharide.

In certain embodiments, the monosaccharide is an N-acetylgalactosamine(GalNAc). GalNAc conjugates, which comprise one or moreN-acetylgalactosamine (GalNAc) derivatives, are described, for example,in U.S. Pat. No. 8,106,022, the entire content of which is herebyincorporated herein by reference. In some embodiments, the GalNAcconjugate serves as a ligand that targets the iRNA to particular cells.In some embodiments, the GalNAc conjugate targets the iRNA to livercells, e.g., by serving as a ligand for the asialoglycoprotein receptorof liver cells (e.g., hepatocytes).

In some embodiments, the carbohydrate conjugate comprises one or moreGalNAc derivatives. The GalNAc derivatives may be attached via a linker,e.g., a bivalent or trivalent branched linker. In some embodiments theGalNAc conjugate is conjugated to the 3′ end of the sense strand. Insome embodiments, the GalNAc conjugate is conjugated to the iRNA agent(e.g., to the 3′ end of the sense strand) via a linker, e.g., a linkeras described herein. In some embodiments the GalNAc conjugate isconjugated to the 5′ end of the sense strand. In some embodiments, theGalNAc conjugate is conjugated to the iRNA agent (e.g., to the 5′ end ofthe sense strand) via a linker, e.g., a linker as described herein.

In certain embodiments of the invention, the GalNAc or GalNAc derivativeis attached to an iRNA agent of the invention via a monovalent linker.In some embodiments, the GalNAc or GalNAc derivative is attached to aniRNA agent of the invention via a bivalent linker. In yet otherembodiments of the invention, the GalNAc or GalNAc derivative isattached to an iRNA agent of the invention via a trivalent linker. Inother embodiments of the invention, the GalNAc or GalNAc derivative isattached to an iRNA agent of the invention via a tetravalent linker.

In certain embodiments, the double stranded RNAi agents of the inventioncomprise one GalNAc or GalNAc derivative attached to the iRNA agent. Incertain embodiments, the double stranded RNAi agents of the inventioncomprise a plurality (e.g., 2, 3, 4, 5, or 6) GalNAc or GalNAcderivatives, each independently attached to a plurality of nucleotidesof the double stranded RNAi agent through a plurality of monovalentlinkers.

In some embodiments, for example, when the two strands of an iRNA agentof the invention are part of one larger molecule connected by anuninterrupted chain of nucleotides between the 3′-end of one strand andthe 5′-end of the respective other strand forming a hairpin loopcomprising, a plurality of unpaired nucleotides, each unpairednucleotide within the hairpin loop may independently comprise a GalNAcor GalNAc derivative attached via a monovalent linker. The hairpin loopmay also be formed by an extended overhang in one strand of the duplex.

In some embodiments, for example, when the two strands of an iRNA agentof the invention are part of one larger molecule connected by anuninterrupted chain of nucleotides between the 3′-end of one strand andthe 5′-end of the respective other strand forming a hairpin loopcomprising, a plurality of unpaired nucleotides, each unpairednucleotide within the hairpin loop may independently comprise a GalNAcor GalNAc derivative attached via a monovalent linker. The hairpin loopmay also be formed by an extended overhang in one strand of the duplex.

In one embodiment, a carbohydrate conjugate for use in the compositionsand methods of the invention is selected from the group consisting of:

wherein Y is O or S and n is 3-6 (Formu

wherein Y is O or S and n is 3-6 (Formula XXV);

wherein X is O or S (Formula XXVII);

In another embodiment, a carbohydrate conjugate for use in thecompositions and methods of the invention is a monosaccharide. In oneembodiment, the monosaccharide is an N-acetylgalactosamine, such as

In some embodiments, the RNAi agent is attached to the carbohydrateconjugate via a linker as shown in the following schematic, wherein X isO or S

In some embodiments, the RNAi agent is conjugated to L96 as defined inTable 1 and shown below:

Another representative carbohydrate conjugate for use in the embodimentsdescribed herein includes, but is not limited to,

(Formula XXXVI), when one of X or Y is an oligonucleotide, the other isa hydrogen.

In some embodiments, a suitable ligand is a ligand disclosed in WO2019/055633, the entire contents of which are incorporated herein byreference. In one embodiment the ligand comprises the structure below:

In certain embodiments of the invention, the GalNAc or GalNAc derivativeis attached to an iRNA agent of the invention via a monovalent linker.In some embodiments, the GalNAc or GalNAc derivative is attached to aniRNA agent of the invention via a bivalent linker. In yet otherembodiments of the invention, the GalNAc or GalNAc derivative isattached to an iRNA agent of the invention via a trivalent linker.

In one embodiment, the double stranded RNAi agents of the inventioncomprise one or more GalNAc or GalNAc derivative attached to the iRNAagent. The GalNAc may be attached to any nucleotide via a linker on thesense strand or antisense strand. The GalNac may be attached to the5′-end of the sense strand, the 3′ end of the sense strand, the 5′-endof the antisense strand, or the 3′-end of the antisense strand. In oneembodiment, the GalNAc is attached to the 3′ end of the sense strand,e.g., via a trivalent linker.

In other embodiments, the double stranded RNAi agents of the inventioncomprise a plurality (e.g., 2, 3, 4, 5, or 6) GalNAc or GalNAcderivatives, each independently attached to a plurality of nucleotidesof the double stranded RNAi agent through a plurality of linkers, e.g.,monovalent linkers.

In some embodiments, for example, when the two strands of an iRNA agentof the invention is part of one larger molecule connected by anuninterrupted chain of nucleotides between the 3′-end of one strand andthe 5′-end of the respective other strand forming a hairpin loopcomprising, a plurality of unpaired nucleotides, each unpairednucleotide within the hairpin loop may independently comprise a GalNAcor GalNAc derivative attached via a monovalent linker.

In some embodiments, the carbohydrate conjugate further comprises one ormore additional ligands as described above, such as, but not limited to,a PK modulator or a cell permeation peptide.

Additional carbohydrate conjugates and linkers suitable for use in thepresent invention include those described in PCT Publication Nos. WO2014/179620 and WO 2014/179627, the entire contents of each of which areincorporated herein by reference.

D. Linkers

In some embodiments, the conjugate or ligand described herein can beattached to an iRNA oligonucleotide with various linkers that can becleavable or non-cleavable.

The term “linker” or “linking group” means an organic moiety thatconnects two parts of a compound, e.g., covalently attaches two parts ofa compound. Linkers typically comprise a direct bond or an atom such asoxygen or sulfur, a unit such as NR8, C(O), C(O)NH, SO, SO₂, SO₂NH or achain of atoms, such as, but not limited to, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, arylalkyl, arylalkenyl, arylalkynyl,heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl,heterocyclylalkyl, heterocyclylalkenyl, heterocyclylalkynyl, aryl,heteroaryl, heterocyclyl, cycloalkyl, cycloalkenyl, alkylarylalkyl,alkylarylalkenyl, alkylarylalkynyl, alkenylarylalkyl,alkenylarylalkenyl, alkenylarylalkynyl, alkynylarylalkyl,alkynylarylalkenyl, alkynylarylalkynyl, alkylheteroarylalkyl,alkylheteroarylalkenyl, alkylheteroarylalkynyl, alkenylheteroarylalkyl,alkenylheteroarylalkenyl, alkenylheteroarylalkynyl,alkynylheteroarylalkyl, alkynylheteroarylalkenyl,alkynylheteroarylalkynyl, alkylheterocyclylalkyl,alkylheterocyclylalkenyl, alkylhererocyclylalkynyl,alkenylheterocyclylalkyl, alkenylheterocyclylalkenyl,alkenylheterocyclylalkynyl, alkynylheterocyclylalkyl,alkynylheterocyclylalkenyl, alkynylheterocyclylalkynyl, alkylaryl,alkenylaryl, alkynylaryl, alkylheteroaryl, alkenylheteroaryl,alkenylheteroaryl, which one or more methylenes can be interrupted orterminated by O, S, S(O), SO₂, N(R8), C(O), substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, or substituted orunsubstituted heterocyclic; where R8 is hydrogen, acyl, aliphatic, orsubstituted aliphatic. In one embodiment, the linker is about 1-24atoms, 2-24, 3-24, 4-24, 5-24, 6-24, 6-18, 7-18, 8-18, 7-17, 8-17, 6-16,7-17, or 8-16 atoms.

A cleavable linking group is one which is sufficiently stable outsidethe cell, but which upon entry into a target cell is cleaved to releasethe two parts the linker is holding together. In one embodiment, thecleavable linking group is cleaved at least about 10 times, 20, times,30 times, 40 times, 50 times, 60 times, 70 times, 80 times, 90 times, ormore, or at least 100 times faster in a target cell or under a firstreference condition (which can, e.g., be selected to mimic or representintracellular conditions) than in the blood of a subject, or under asecond reference condition (which can, e.g., be selected to mimic orrepresent conditions found in the blood or serum).

Cleavable linking groups are susceptible to cleavage agents, e.g., pH,redox potential, or the presence of degradative molecules. Generally,cleavage agents are more prevalent or found at higher levels oractivities inside cells than in serum or blood. Examples of suchdegradative agents include: redox agents which are selected forparticular substrates or which have no substrate specificity, including,e.g., oxidative or reductive enzymes or reductive agents such asmercaptans, present in cells, that can degrade a redox cleavable linkinggroup by reduction; esterases; endosomes or agents that can create anacidic environment, e.g., those that result in a pH of five or lower;enzymes that can hydrolyze or degrade an acid cleavable linking group byacting as a general acid, peptidases (which can be substrate specific),and phosphatases.

A cleavable linkage group, such as a disulfide bond can be susceptibleto pH. The pH of human serum is 7.4, while the average intracellular pHis slightly lower, ranging from about 7.1-7.3. Endosomes have a moreacidic pH, in the range of 5.5-6.0, and lysosomes have an even moreacidic pH at around 5.0. Some linkers will have a cleavable linkinggroup that is cleaved at a selected pH, thereby releasing a cationiclipid from the ligand inside the cell, or into the desired compartmentof the cell.

A linker can include a cleavable linking group that is cleavable by aparticular enzyme. The type of cleavable linking group incorporated intoa linker can depend on the cell to be targeted. For example, aliver-targeting ligand can be linked to a cationic lipid through alinker that includes an ester group. Liver cells are rich in esterases,and therefore the linker will be cleaved more efficiently in liver cellsthan in cell types that are not esterase-rich. Other cell-types rich inesterases include cells of the lung, renal cortex, and testis.

Linkers that contain peptide bonds can be used when targeting cell typesrich in peptidases, such as liver cells and synoviocytes.

In general, the suitability of a candidate cleavable linking group canbe evaluated by testing the ability of a degradative agent (orcondition) to cleave the candidate linking group. It will also bedesirable to also test the candidate cleavable linking group for theability to resist cleavage in the blood or when in contact with othernon-target tissue. Thus, one can determine the relative susceptibilityto cleavage between a first and a second condition, where the first isselected to be indicative of cleavage in a target cell and the second isselected to be indicative of cleavage in other tissues or biologicalfluids, e.g., blood or serum. The evaluations can be carried out in cellfree systems, in cells, in cell culture, in organ or tissue culture, orin whole animals. It can be useful to make initial evaluations incell-free or culture conditions and to confirm by further evaluations inwhole animals. In certain embodiments, useful candidate compounds arecleaved at least about 2, 4, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100times faster in the cell (or under in vitro conditions selected to mimicintracellular conditions) as compared to blood or serum (or under invitro conditions selected to mimic extracellular conditions).

i. Redox Cleavable Linking Groups

In certain embodiments, a cleavable linking group is a redox cleavablelinking group that is cleaved upon reduction or oxidation. An example ofreductively cleavable linking group is a disulphide linking group(—S—S—). To determine if a candidate cleavable linking group is asuitable “reductively cleavable linking group,” or for example issuitable for use with a particular iRNA moiety and particular targetingagent one can look to methods described herein. For example, a candidatecan be evaluated by incubation with dithiothreitol (DTT), or otherreducing agent using reagents know in the art, which mimic the rate ofcleavage which would be observed in a cell, e.g., a target cell. Thecandidates can also be evaluated under conditions which are selected tomimic blood or serum conditions. In one, candidate compounds are cleavedby at most about 10% in the blood. In other embodiments, usefulcandidate compounds are degraded at least about 2, 4, 10, 20, 30, 40,50, 60, 70, 80, 90, or about 100 times faster in the cell (or under invitro conditions selected to mimic intracellular conditions) as comparedto blood (or under in vitro conditions selected to mimic extracellularconditions). The rate of cleavage of candidate compounds can bedetermined using standard enzyme kinetics assays under conditions chosento mimic intracellular media and compared to conditions chosen to mimicextracellular media.

ii. Phosphate-Based Cleavable Linking Groups

In other embodiments, a cleavable linker comprises a phosphate-basedcleavable linking group. A phosphate-based cleavable linking group iscleaved by agents that degrade or hydrolyze the phosphate group. Anexample of an agent that cleaves phosphate groups in cells are enzymessuch as phosphatases in cells. Examples of phosphate-based linkinggroups are —O—P(O)(ORk)-O—, —O—P(S)(ORk)-O—, —O—P(S)(SRk)-O—,—S—P(O)(ORk)-O—, —O—P(O)(ORk)-S—, —S—P(O)(ORk)-S—, —O—P(S)(ORk)-S—,—S—P(S)(ORk)-O—, —O—P(O)(Rk)-O—, —O—P(S)(Rk)-O—, —S—P(O)(Rk)-O—,—S—P(S)(Rk)-O—, —S—P(O)(Rk)-S—, —O—P(S)(Rk)-S—, wherein Rk at eachoccurrence can be, independently, C1-C20 alkyl, C1-C20 haloalkyl, C6-C10aryl, or C7-C12 aralkyl. Exemplary embodiments include —O—P(O)(OH)—O—,—O—P(S)(OH)—O—, —O—P(S)(SH)—O—, —S—P(O)(OH)—O—, —O—P(O)(OH)—S—,—S—P(O)(OH)—S—, —O—P(S)(OH)—S—, —S—P(S)(OH)—O—, —O—P(O)(H)—O—,—O—P(S)(H)—O—, —S—P(O)(H)—O, —S—P(S)(H)—O—, —S—P(O)(H)—S—, and—O—P(S)(H)—S—. In certain embodiments, a phosphate-based linking groupis —O—P(O)(OH)—O—. These candidates can be evaluated using methodsanalogous to those described above.

iii. Acid Cleavable Linking Groups

In other embodiments, a cleavable linker comprises an acid cleavablelinking group. An acid cleavable linking group is a linking group thatis cleaved under acidic conditions. In certain embodiments acidcleavable linking groups are cleaved in an acidic environment with a pHof about 6.5 or lower (e.g., about 6.0, 5.5, 5.0, or lower), or byagents such as enzymes that can act as a general acid. In a cell,specific low pH organelles, such as endosomes and lysosomes can providea cleaving environment for acid cleavable linking groups. Examples ofacid cleavable linking groups include but are not limited to hydrazones,esters, and esters of amino acids. Acid cleavable groups can have thegeneral formula —C═NN—, C(O)O, or —OC(O). An exemplary embodiment iswhen the carbon attached to the oxygen of the ester (the alkoxy group)is an aryl group, substituted alkyl group, or tertiary alkyl group suchas dimethyl pentyl or t-butyl. These candidates can be evaluated usingmethods analogous to those described above.

iv. Ester-Based Linking Groups

In other embodiments, a cleavable linker comprises an ester-basedcleavable linking group. An ester-based cleavable linking group iscleaved by enzymes such as esterases and amidases in cells. Examples ofester-based cleavable linking groups include, but are not limited to,esters of alkylene, alkenylene and alkynylene groups. Ester cleavablelinking groups have the general formula —C(O)O—, or —OC(O)—. Thesecandidates can be evaluated using methods analogous to those describedabove.

v. Peptide-Based Cleaving Groups

In yet other embodiments, a cleavable linker comprises a peptide-basedcleavable linking group. A peptide-based cleavable linking group iscleaved by enzymes such as peptidases and proteases in cells.Peptide-based cleavable linking groups are peptide bonds formed betweenamino acids to yield oligopeptides (e.g., dipeptides, tripeptides etc.)and polypeptides. Peptide-based cleavable groups do not include theamide group (—C(O)NH—). The amide group can be formed between anyalkylene, alkenylene or alkynelene. A peptide bond is a special type ofamide bond formed between amino acids to yield peptides and proteins.The peptide based cleavage group is generally limited to the peptidebond (i.e., the amide bond) formed between amino acids yielding peptidesand proteins and does not include the entire amide functional group.Peptide-based cleavable linking groups have the general formula—NHCHRAC(O)NHCHRBC(O)—, where RA and RB are the R groups of the twoadjacent amino acids. These candidates can be evaluated using methodsanalogous to those described above.

In some embodiments, an iRNA of the invention is conjugated to acarbohydrate through a linker. Non-limiting examples of iRNAcarbohydrate conjugates with linkers of the compositions and methods ofthe invention include, but are not limited to,

when one of X or Y is an oligonucleotide, the other is a hydrogen.

In certain embodiments of the compositions and methods of the invention,a ligand is one or more “GalNAc” (N-acetylgalactosamine) derivativesattached through a bivalent or trivalent branched linker.

In one embodiment, a dsRNA of the invention is conjugated to a bivalentor trivalent branched linker selected from the group of structures shownin any of formula (XLV)-(XLVIII):

wherein:q2A, q2B, q3A, q3B, q4A, q4B, q5A, q5B and q5C represent independentlyfor each occurrence 0-20 and wherein the repeating unit can be the sameor different;P^(2A), P^(2B), P^(3A), P^(3B), P^(4A), P^(4B), P^(5A), P^(5B), P^(5C),T^(2A), T^(2B), T^(3A), T^(3B), T^(4A), T^(4B), T^(4A), T^(5B), T^(5C)are each independently for each occurrence absent, CO, NH, O, S, OC(O),NHC(O), CH₂, CH₂NH or CH₂O;Q^(2A), Q^(2B), Q^(3A), Q^(3B), Q^(4A), Q^(4B), Q^(5A), Q^(5B), Q^(5C)are independently for each occurrence absent, alkylene, substitutedalkylene wherein one or more methylenes can be interrupted or terminatedby one or more of O, S, S(O), SO₂, N(R^(N)), C(R′)═C(R″), C≡C or C(O);R^(2A), R^(2B), R^(3A), R^(3B), R^(4A), R^(4B), R^(5A), R^(5B), R^(5C)are each independently for each occurrence absent, NH, O, S, CH₂, C(O)O,C(O)NH, NHCH(R^(a))C(O), —C(O)—CH(R^(a))—NH—, CO, CH═N—O,

or heterocyclyl; L^(2A), L^(2B), L^(3A), L^(3B), L^(4A), L^(4B), L^(5A),L^(5B) and L^(5C) represent the ligand; i.e. each independently for eachoccurrence a monosaccharide (such as GalNAc), disaccharide,trisaccharide, tetrasaccharide, oligosaccharide, or polysaccharide; andR^(a) is H or amino acid side chain. Trivalent conjugating GalNAcderivatives are particularly useful for use with RNAi agents forinhibiting the expression of a target gene, such as those of formula(XLIX):

wherein L^(5A), L^(5B) and L^(5C) represent a monosaccharide, such asGalNAc derivative.

Examples of suitable bivalent and trivalent branched linker groupsconjugating GalNAc derivatives include, but are not limited to, thestructures recited above as formulas II, VII, XI, X, and XIII.

Representative U.S. patents that teach the preparation of RNA conjugatesinclude, but are not limited to, U.S. Pat. Nos. 4,828,979; 4,948,882;5,218,105; 5,525,465; 5,541,313; 5,545,730; 5,552,538; 5,578,717,5,580,731; 5,591,584; 5,109,124; 5,118,802; 5,138,045; 5,414,077;5,486,603; 5,512,439; 5,578,718; 5,608,046; 4,587,044; 4,605,735;4,667,025; 4,762,779; 4,789,737; 4,824,941; 4,835,263; 4,876,335;4,904,582; 4,958,013; 5,082,830; 5,112,963; 5,214,136; 5,082,830;5,112,963; 5,214,136; 5,245,022; 5,254,469; 5,258,506; 5,262,536;5,272,250; 5,292,873; 5,317,098; 5,371,241, 5,391,723; 5,416,203,5,451,463; 5,510,475; 5,512,667; 5,514,785; 5,565,552; 5,567,810;5,574,142; 5,585,481; 5,587,371; 5,595,726; 5,597,696; 5,599,923;5,599,928; 5,688,941; 6,294,664; 6,320,017; 6,576,752; 6,783,931;6,900,297; 7,037,646; and 8,106,022, the entire contents of each ofwhich are hereby incorporated herein by reference.

It is not necessary for all positions in a given compound to beuniformly modified, and in fact more than one of the aforementionedmodifications can be incorporated in a single compound or even at asingle nucleoside within an iRNA. The present invention also includesiRNA compounds that are chimeric compounds.

“Chimeric” iRNA compounds or “chimeras,” in the context of thisinvention, are iRNA compounds, such as, dsRNAi agents, that contain twoor more chemically distinct regions, each made up of at least onemonomer unit, i.e., a nucleotide in the case of a dsRNA compound. TheseiRNAs typically contain at least one region wherein the RNA is modifiedso as to confer upon the iRNA increased resistance to nucleasedegradation, increased cellular uptake, or increased binding affinityfor the target nucleic acid. An additional region of the iRNA can serveas a substrate for enzymes capable of cleaving RNA:DNA or RNA:RNAhybrids. By way of example, RNase H is a cellular endonuclease whichcleaves the RNA strand of an RNA:DNA duplex. Activation of RNase H,therefore, results in cleavage of the RNA target, thereby greatlyenhancing the efficiency of iRNA inhibition of gene expression.Consequently, comparable results can often be obtained with shorteriRNAs when chimeric dsRNAs are used, compared to phosphorothioate deoxydsRNAs hybridizing to the same target region. Cleavage of the RNA targetcan be routinely detected by gel electrophoresis and, if necessary,associated nucleic acid hybridization techniques known in the art.

In certain instances, the RNA of an iRNA can be modified by a non-ligandgroup. A number of non-ligand molecules have been conjugated to iRNAs inorder to enhance the activity, cellular distribution or cellular uptakeof the iRNA, and procedures for performing such conjugations areavailable in the scientific literature. Such non-ligand moieties haveincluded lipid moieties, such as cholesterol (Kubo, T. et al., Biochem.Biophys. Res. Comm., 2007, 365(1):54-61; Letsinger et al., Proc. Natl.Acad. Sci. USA, 1989, 86:6553), cholic acid (Manoharan et al., Bioorg.Med. Chem. Lett., 1994, 4:1053), a thioether, e.g., hexyl-S-tritylthiol(Manoharan et al., Ann. N.Y. Acad. Sci., 1992, 660:306; Manoharan etal., Bioorg. Med. Chem. Let., 1993, 3:2765), a thiocholesterol(Oberhauser et al., Nucl. Acids Res., 1992, 20:533), an aliphatic chain,e.g., dodecandiol or undecyl residues (Saison-Behmoaras et al., EMBO J.,1991, 10:111; Kabanov et al., FEBS Lett., 1990, 259:327; Svinarchuk etal., Biochimie, 1993, 75:49), a phospholipid, e.g.,di-hexadecyl-rac-glycerol or triethylammonium1,2-di-O-hexadecyl-rac-glycero-3-H-phosphonate (Manoharan et al.,Tetrahedron Lett., 1995, 36:3651; Shea et al., Nucl. Acids Res., 1990,18:3777), a polyamine or a polyethylene glycol chain (Manoharan et al.,Nucleosides & Nucleotides, 1995, 14:969), or adamantane acetic acid(Manoharan et al., Tetrahedron Lett., 1995, 36:3651), a palmityl moiety(Mishra et al., Biochim. Biophys. Acta, 1995, 1264:229), or anoctadecylamine or hexylamino-carbonyl-oxycholesterol moiety (Crooke etal., J. Pharmacol. Exp. Ther., 1996, 277:923). Representative UnitedStates patents that teach the preparation of such RNA conjugates havebeen listed above. Typical conjugation protocols involve the synthesisof RNAs bearing an aminolinker at one or more positions of the sequence.The amino group is then reacted with the molecule being conjugated usingappropriate coupling or activating reagents. The conjugation reactioncan be performed either with the RNA still bound to the solid support orfollowing cleavage of the RNA, in solution phase. Purification of theRNA conjugate by HPLC typically affords the pure conjugate.

IV. Delivery of an iRNA of the Invention

The delivery of an iRNA of the invention to a cell e.g., a cell within asubject, such as a human subject (e.g., a subject in need thereof, suchas a subject susceptible to or diagnosed with a complement factorB-associated disorder) can be achieved in a number of different ways.For example, delivery may be performed by contacting a cell with an iRNAof the invention either in vitro or in vivo. In vivo delivery may alsobe performed directly by administering a composition comprising an iRNA,e.g., a dsRNA, to a subject. Alternatively, in vivo delivery may beperformed indirectly by administering one or more vectors that encodeand direct the expression of the iRNA. These alternatives are discussedfurther below.

In general, any method of delivering a nucleic acid molecule (in vitroor in vivo) can be adapted for use with an iRNA of the invention (seee.g., Akhtar S. and Julian R L. (1992) Trends Cell. Biol. 2(5):139-144and WO94/02595, which are incorporated herein by reference in theirentireties). For in vivo delivery, factors to consider in order todeliver an iRNA molecule include, for example, biological stability ofthe delivered molecule, prevention of non-specific effects, andaccumulation of the delivered molecule in the target tissue. RNAinterference has also shown success with local delivery to the CNS bydirect injection (Dorn, G., et al. (2004) Nucleic Acids 32:e49; Tan, PH., et al (2005) Gene Ther. 12:59-66; Makimura, H., et al (2002) BMCNeurosci. 3:18; Shishkina, G T., et al (2004) Neuroscience 129:521-528;Thakker, E R., et al (2004) Proc. Natl. Acad. Sci. U.S.A.101:17270-17275; Akaneya, Y., et al (2005) J. Neurophysiol. 93:594-602).Modification of the RNA or the pharmaceutical carrier can also permittargeting of the iRNA to the target tissue and avoid undesirableoff-target effects. iRNA molecules can be modified by chemicalconjugation to lipophilic groups such as cholesterol to enhance cellularuptake and prevent degradation. For example, an iRNA directed againstApoB conjugated to a lipophilic cholesterol moiety was injectedsystemically into mice and resulted in knockdown of apoB mRNA in boththe liver and jejunum (Soutschek, J., et al (2004) Nature 432:173-178).

In an alternative embodiment, the iRNA can be delivered using drugdelivery systems such as a nanoparticle, a dendrimer, a polymer,liposomes, or a cationic delivery system. Positively charged cationicdelivery systems facilitate binding of an iRNA molecule (negativelycharged) and also enhance interactions at the negatively charged cellmembrane to permit efficient uptake of an iRNA by the cell. Cationiclipids, dendrimers, or polymers can either be bound to an iRNA, orinduced to form a vesicle or micelle (see e.g., Kim S H, et al (2008)Journal of Controlled Release 129(2):107-116) that encases an iRNA. Theformation of vesicles or micelles further prevents degradation of theiRNA when administered systemically. Methods for making andadministering cationic—iRNA complexes are well within the abilities ofone skilled in the art (see e.g., Sorensen, D R, et al (2003) J. Mol.Biol 327:761-766; Verma, U N, et al (2003) Clin. Cancer Res.9:1291-1300; Arnold, A S et al (2007) J. Hypertens. 25:197-205, whichare incorporated herein by reference in their entirety). Somenon-limiting examples of drug delivery systems useful for systemicdelivery of iRNAs include DOTAP (Sorensen, D R., et al (2003), supra;Verma, U N, et al (2003), supra), “solid nucleic acid lipid particles”(Zimmermann, T S, et al (2006) Nature 441:111-114), cardiolipin (Chien,P Y, et al (2005) Cancer Gene Ther. 12:321-328; Pal, A, et al (2005) IntJ. Oncol. 26:1087-1091), polyethyleneimine (Bonnet M E, et al (2008)Pharm. Res. August 16 Epub ahead of print; Aigner, A. (2006) J. Biomed.Biotechnol. 71659), Arg-Gly-Asp (RGD) peptides (Liu, S. (2006) Mol.Pharm. 3:472-487), and polyamidoamines (Tomalia, D A, et al (2007)Biochem. Soc. Trans. 35:61-67; Yoo, H., et al (1999) Pharm. Res.16:1799-1804). In some embodiments, an iRNA forms a complex withcyclodextrin for systemic administration. Methods for administration andpharmaceutical compositions of iRNAs and cyclodextrins can be found inU.S. Pat. No. 7,427,605, which is herein incorporated by reference inits entirety.

A. Vector Encoded iRNAs of the Invention

iRNA targeting the complement factor B gene can be expressed fromtranscription units inserted into DNA or RNA vectors (see, e.g.,Couture, A, et al., TIG. (1996), 12:5-10; Skillern, A, et al.,International PCT Publication No. WO 00/22113, Conrad, International PCTPublication No. WO 00/22114, and Conrad, U.S. Pat. No. 6,054,299).Expression can be transient (on the order of hours to weeks) orsustained (weeks to months or longer), depending upon the specificconstruct used and the target tissue or cell type. These transgenes canbe introduced as a linear construct, a circular plasmid, or a viralvector, which can be an integrating or non-integrating vector. Thetransgene can also be constructed to permit it to be inherited as anextrachromosomal plasmid (Gassmann, et al., Proc. Natl. Acad. Sci. USA(1995) 92:1292).

Viral vector systems which can be utilized with the methods andcompositions described herein include, but are not limited to, (a)adenovirus vectors; (b) retrovirus vectors, including but not limited tolentiviral vectors, moloney murine leukemia virus, etc.; (c)adeno-associated virus vectors; (d) herpes simplex virus vectors; (e) SV40 vectors; (f) polyoma virus vectors; (g) papilloma virus vectors; (h)picornavirus vectors; (i) pox virus vectors such as an orthopox, e.g.,vaccinia virus vectors or avipox, e.g. canary pox or fowl pox; and (j) ahelper-dependent or gutless adenovirus. Replication-defective virusescan also be advantageous. Different vectors will or will not becomeincorporated into the cells' genome. The constructs can include viralsequences for transfection, if desired. Alternatively, the construct canbe incorporated into vectors capable of episomal replication, e.g. EPVand EBV vectors. Constructs for the recombinant expression of an iRNAwill generally require regulatory elements, e.g., promoters, enhancers,etc., to ensure the expression of the iRNA in target cells. Otheraspects to consider for vectors and constructs are known in the art.

V. Pharmaceutical Compositions of the Invention

The present invention also includes pharmaceutical compositions andformulations which include the iRNAs of the invention. In oneembodiment, provided herein are pharmaceutical compositions containingan iRNA, as described herein, and a pharmaceutically acceptable carrier.The pharmaceutical compositions containing the iRNA are useful forpreventing or treating a complement factor B-associated disorder. Suchpharmaceutical compositions are formulated based on the mode ofdelivery. One example is compositions that are formulated for systemicadministration via parenteral delivery, e.g., by subcutaneous (SC),intramuscular (IM), or intravenous (IV) delivery. The pharmaceuticalcompositions of the invention may be administered in dosages sufficientto inhibit expression of a complement factor B gene.

In some embodiments, the pharmaceutical compositions of the inventionare sterile. In another embodiment, the pharmaceutical compositions ofthe invention are pyrogen free.

The pharmaceutical compositions of the invention may be administered indosages sufficient to inhibit expression of a complement factor B gene.In general, a suitable dose of an iRNA of the invention will be in therange of about 0.001 to about 200.0 milligrams per kilogram body weightof the recipient per day, generally in the range of about 1 to 50 mg perkilogram body weight per day. Typically, a suitable dose of an iRNA ofthe invention will be in the range of about 0.1 mg/kg to about 5.0mg/kg, such as, about 0.3 mg/kg and about 3.0 mg/kg. A repeat-doseregimen may include administration of a therapeutic amount of iRNA on aregular basis, such as every month, once every 3-6 months, or once ayear. In certain embodiments, the iRNA is administered about once permonth to about once per six months.

After an initial treatment regimen, the treatments can be administeredon a less frequent basis. Duration of treatment can be determined basedon the severity of disease.

In other embodiments, a single dose of the pharmaceutical compositionscan be long lasting, such that doses are administered at not more than1, 2, 3, or 4 month intervals. In some embodiments of the invention, asingle dose of the pharmaceutical compositions of the invention isadministered about once per month. In other embodiments of theinvention, a single dose of the pharmaceutical compositions of theinvention is administered quarterly (i.e., about every three months). Inother embodiments of the invention, a single dose of the pharmaceuticalcompositions of the invention is administered twice per year (i.e.,about once every six months).

The skilled artisan will appreciate that certain factors can influencethe dosage and timing required to effectively treat a subject, includingbut not limited to mutations present in the subject, previoustreatments, the general health or age of the subject, and other diseasespresent. Moreover, treatment of a subject with a prophylactically ortherapeutically effective amount, as appropriate, of a composition caninclude a single treatment or a series of treatments.

The iRNA can be delivered in a manner to target a particular tissue(e.g., hepatocytes).

Pharmaceutical compositions of the present invention include, but arenot limited to, solutions, emulsions, and liposome-containingformulations. These compositions can be generated from a variety ofcomponents that include, but are not limited to, preformed liquids,self-emulsifying solids, and self-emulsifying semisolids. Formulationsinclude those that target the liver.

The pharmaceutical formulations of the present invention, which canconveniently be presented in unit dosage form, can be prepared accordingto conventional techniques well known in the pharmaceutical industry.Such techniques include the step of bringing into association the activeingredients with the pharmaceutical carrier(s) or excipient(s). Ingeneral, the formulations are prepared by uniformly and intimatelybringing into association the active ingredients with liquid carriers.

A. Additional Formulations

i. Emulsions

The compositions of the present invention can be prepared and formulatedas emulsions.

Emulsions are typically heterogeneous systems of one liquid dispersed inanother in the form of droplets usually exceeding 0.1 μm in diameter(see e.g., Ansel's Pharmaceutical Dosage Forms and Drug DeliverySystems, Allen, L V., Popovich N G., and Ansel H C., 2004, LippincottWilliams & Wilkins (8th ed.), New York, N.Y.; Idson, in PharmaceuticalDosage Forms, Lieberman, Rieger and Banker (Eds.), 1988, Marcel Dekker,Inc., New York, N.Y., volume 1, p. 199; Rosoff, in Pharmaceutical DosageForms, Lieberman, Rieger and Banker (Eds.), 1988, Marcel Dekker, Inc.,New York, N.Y., Volume 1, p. 245; Block in Pharmaceutical Dosage Forms,Lieberman, Rieger and Banker (Eds.), 1988, Marcel Dekker, Inc., NewYork, N.Y., volume 2, p. 335; Higuchi et al., in Remington'sPharmaceutical Sciences, Mack Publishing Co., Easton, Pa., 1985, p.301). Emulsions are often biphasic systems comprising two immiscibleliquid phases intimately mixed and dispersed with each other. Ingeneral, emulsions can be of either the water-in-oil (w/o) or theoil-in-water (o/w) variety. When an aqueous phase is finely divided intoand dispersed as minute droplets into a bulk oily phase, the resultingcomposition is called a water-in-oil (w/o) emulsion. Alternatively, whenan oily phase is finely divided into and dispersed as minute dropletsinto a bulk aqueous phase, the resulting composition is called anoil-in-water (o/w) emulsion. Emulsions can contain additional componentsin addition to the dispersed phases, and the active drug which can bepresent as a solution either in the aqueous phase, oily phase or itselfas a separate phase. Pharmaceutical excipients such as emulsifiers,stabilizers, dyes, and anti-oxidants can also be present in emulsions asneeded. Pharmaceutical emulsions can also be multiple emulsions that arecomprised of more than two phases such as, for example, in the case ofoil-in-water-in-oil (o/w/o) and water-in-oil-in-water (w/o/w) emulsions.Such complex formulations often provide certain advantages that simplebinary emulsions do not. Multiple emulsions in which individual oildroplets of an o/w emulsion enclose small water droplets constitute aw/o/w emulsion. Likewise a system of oil droplets enclosed in globulesof water stabilized in an oily continuous phase provides an o/w/oemulsion.

Emulsions are characterized by little or no thermodynamic stability.Often, the dispersed or discontinuous phase of the emulsion is welldispersed into the external or continuous phase and maintained in thisform through the means of emulsifiers or the viscosity of theformulation. Other means of stabilizing emulsions entail the use ofemulsifiers that can be incorporated into either phase of the emulsion.Emulsifiers can broadly be classified into four categories: syntheticsurfactants, naturally occurring emulsifiers, absorption bases, andfinely dispersed solids (see e.g., Ansel's Pharmaceutical Dosage Formsand Drug Delivery Systems, Allen, L V., Popovich N G., and Ansel H C.,2004, Lippincott Williams & Wilkins (8th ed.), New York, N.Y.; Idson, inPharmaceutical Dosage Forms, Lieberman, Rieger and Banker (Eds.), 1988,Marcel Dekker, Inc., New York, N.Y., volume 1, p. 199).

Synthetic surfactants, also known as surface active agents, have foundwide applicability in the formulation of emulsions and have beenreviewed in the literature (see e.g., Ansel's Pharmaceutical DosageForms and Drug Delivery Systems, Allen, L V., Popovich N G., and Ansel HC., 2004, Lippincott Williams & Wilkins (8th ed.), New York, N.Y.;Rieger, in Pharmaceutical Dosage Forms, Lieberman, Rieger and Banker(Eds.), 1988, Marcel Dekker, Inc., New York, N.Y., volume 1, p. 285;Idson, in Pharmaceutical Dosage Forms, Lieberman, Rieger and Banker(Eds.), Marcel Dekker, Inc., New York, N.Y., 1988, volume 1, p. 199).Surfactants are typically amphiphilic and comprise a hydrophilic and ahydrophobic portion. The ratio of the hydrophilic to the hydrophobicnature of the surfactant has been termed the hydrophile/lipophilebalance (HLB) and is a valuable tool in categorizing and selectingsurfactants in the preparation of formulations. Surfactants can beclassified into different classes based on the nature of the hydrophilicgroup: nonionic, anionic, cationic, and amphoteric (see e.g., Ansel'sPharmaceutical Dosage Forms and Drug Delivery Systems, Allen, L V.,Popovich N G., and Ansel H C., 2004, Lippincott Williams & Wilkins (8thed.), New York, N.Y. Rieger, in Pharmaceutical Dosage Forms, Lieberman,Rieger and Banker (Eds.), 1988, Marcel Dekker, Inc., New York, N.Y.,volume 1, p. 285).

A large variety of non-emulsifying materials are also included inemulsion formulations and contribute to the properties of emulsions.These include fats, oils, waxes, fatty acids, fatty alcohols, fattyesters, humectants, hydrophilic colloids, preservatives, andantioxidants (Block, in Pharmaceutical Dosage Forms, Lieberman, Riegerand Banker (Eds.), 1988, Marcel Dekker, Inc., New York, N.Y., volume 1,p. 335; Idson, in Pharmaceutical Dosage Forms, Lieberman, Rieger andBanker (Eds.), 1988, Marcel Dekker, Inc., New York, N.Y., volume 1, p.199).

The application of emulsion formulations via dermatological, oral, andparenteral routes, and methods for their manufacture have been reviewedin the literature (see e.g., Ansel's Pharmaceutical Dosage Forms andDrug Delivery Systems, Allen, L V., Popovich N G., and Ansel H C., 2004,Lippincott Williams & Wilkins (8th ed.), New York, N.Y.; Idson, inPharmaceutical Dosage Forms, Lieberman, Rieger and Banker (Eds.), 1988,Marcel Dekker, Inc., New York, N.Y., volume 1, p. 199).

ii. Microemulsions

In one embodiment of the present invention, the compositions of iRNAsand nucleic acids are formulated as microemulsions. A microemulsion canbe defined as a system of water, oil, and amphiphile which is a singleoptically isotropic and thermodynamically stable liquid solution (seee.g., Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems,Allen, L V., Popovich N G., and Ansel H C., 2004, Lippincott Williams &Wilkins (8th ed.), New York, N.Y.; Rosoff, in Pharmaceutical DosageForms, Lieberman, Rieger and Banker (Eds.), 1988, Marcel Dekker, Inc.,New York, N.Y., volume 1, p. 245). Typically microemulsions are systemsthat are prepared by first dispersing an oil in an aqueous surfactantsolution and then adding a sufficient amount of a fourth component,generally an intermediate chain-length alcohol to form a transparentsystem. Therefore, microemulsions have also been described asthermodynamically stable, isotropically clear dispersions of twoimmiscible liquids that are stabilized by interfacial films ofsurface-active molecules (Leung and Shah, in: Controlled Release ofDrugs: Polymers and Aggregate Systems, Rosoff, M., Ed., 1989, VCHPublishers, New York, pages 185-215).

iii. Microparticles

An iRNA of the invention may be incorporated into a particle, e.g., amicroparticle. Microparticles can be produced by spray-drying, but mayalso be produced by other methods including lyophilization, evaporation,fluid bed drying, vacuum drying, or a combination of these techniques.

iv. Penetration Enhancers

In one embodiment, the present invention employs various penetrationenhancers to effect the efficient delivery of nucleic acids,particularly iRNAs, to the skin of animals. Most drugs are present insolution in both ionized and nonionized forms. However, usually onlylipid soluble or lipophilic drugs readily cross cell membranes. It hasbeen discovered that even non-lipophilic drugs can cross cell membranesif the membrane to be crossed is treated with a penetration enhancer. Inaddition to aiding the diffusion of non-lipophilic drugs across cellmembranes, penetration enhancers also enhance the permeability oflipophilic drugs.

Penetration enhancers can be classified as belonging to one of fivebroad categories, i.e., surfactants, fatty acids, bile salts, chelatingagents, and non-chelating non-surfactants (see e.g., Malmsten, M.Surfactants and polymers in drug delivery, Informa Health Care, NewYork, N.Y., 2002; Lee et al., Critical Reviews in Therapeutic DrugCarrier Systems, 1991, p. 92). Each of the above mentioned classes ofpenetration enhancers and their use in manufacture of pharmaceuticalcompositions and delivery of pharmaceutical agents are well known in theart.

v. Excipients

In contrast to a carrier compound, a “pharmaceutical carrier” or“excipient” is a pharmaceutically acceptable solvent, suspending agent,or any other pharmacologically inert vehicle for delivering one or morenucleic acids to an animal. The excipient can be liquid or solid and isselected, with the planned manner of administration in mind, so as toprovide for the desired bulk, consistency, etc., when combined with anucleic acid and the other components of a given pharmaceuticalcomposition. Such agent are well known in the art.

vi. Other Components

The compositions of the present invention can additionally contain otheradjunct components conventionally found in pharmaceutical compositions,at their art-established usage levels. Thus, for example, thecompositions can contain additional, compatible, pharmaceutically-activematerials such as, for example, antipruritics, astringents, localanesthetics or anti-inflammatory agents, or can contain additionalmaterials useful in physically formulating various dosage forms of thecompositions of the present invention, such as dyes, flavoring agents,preservatives, antioxidants, opacifiers, thickening agents andstabilizers. However, such materials, when added, should not undulyinterfere with the biological activities of the components of thecompositions of the present invention. The formulations can besterilized and, if desired, mixed with auxiliary agents, e.g.,lubricants, preservatives, stabilizers, wetting agents, emulsifiers,salts for influencing osmotic pressure, buffers, colorings, flavorings,or aromatic substances, and the like which do not deleteriously interactwith the nucleic acid(s) of the formulation.

Aqueous suspensions can contain substances which increase the viscosityof the suspension including, for example, sodium carboxymethylcellulose,sorbitol, or dextran. The suspension can also contain stabilizers.

In some embodiments, pharmaceutical compositions featured in theinvention include (a) one or more iRNA and (b) one or more agents whichfunction by a non-iRNA mechanism and which are useful in treating acomplement factor B-associated disorder.

Toxicity and prophylactic efficacy of such compounds can be determinedby standard pharmaceutical procedures in cell cultures or experimentalanimals, e.g., for determining the LD50 (the dose lethal to 50% of thepopulation) and the ED50 (the dose prophylactically effective in 50% ofthe population). The dose ratio between toxic and therapeutic effects isthe therapeutic index and it can be expressed as the ratio LD50/ED50.Compounds that exhibit high therapeutic indices are preferred.

The data obtained from cell culture assays and animal studies can beused in formulating a range of dosage for use in humans. The dosage ofcompositions featured herein in the invention lies generally within arange of circulating concentrations that include the ED50, such as anED80 or ED90, with little or no toxicity. The dosage can vary withinthis range depending upon the dosage form employed and the route ofadministration utilized. For any compound used in the methods featuredin the invention, the prophylactically effective dose can be estimatedinitially from cell culture assays. A dose can be formulated in animalmodels to achieve a circulating plasma concentration range of thecompound or, when appropriate, of the polypeptide product of a targetsequence (e.g., achieving a decreased concentration of the polypeptide)that includes the IC50 (i.e., the concentration of the test compoundwhich achieves a half-maximal inhibition of symptoms) or higher levelsof inhibition as determined in cell culture. Such information can beused to more accurately determine useful doses in humans. Levels inplasma can be measured, for example, by high performance liquidchromatography.

In addition to their administration, as discussed above, the iRNAsfeatured in the invention can be administered in combination with otherknown agents used for the prevention or treatment of a complement factorB-associated disorder. In any event, the administering physician canadjust the amount and timing of iRNA administration on the basis ofresults observed using standard measures of efficacy known in the art ordescribed herein.

VI. Methods for Inhibiting Complement Factor B Expression

The present invention also provides methods of inhibiting expression ofa CFB gene in a cell. The methods include contacting a cell with an RNAiagent, e.g., double stranded RNA agent, in an amount effective toinhibit expression of CFB in the cell, thereby inhibiting expression ofCFB in the cell.

Contacting of a cell with an iRNA, e.g., a double stranded RNA agent,may be done in vitro or in vivo. Contacting a cell in vivo with the iRNAincludes contacting a cell or group of cells within a subject, e.g., ahuman subject, with the iRNA. Combinations of in vitro and in vivomethods of contacting a cell are also possible. Contacting a cell may bedirect or indirect, as discussed above. Furthermore, contacting a cellmay be accomplished via a targeting ligand, including any liganddescribed herein or known in the art. In certain embodiments, thetargeting ligand is a carbohydrate moiety, e.g., a GalNAc ligand, or anyother ligand that directs the RNAi agent to a site of interest.

The term “inhibiting,” as used herein, is used interchangeably with“reducing,” “silencing,” “downregulating”, “suppressing”, and othersimilar terms, and includes any level of inhibition.

The phrase “inhibiting expression of a complement factor B gene” isintended to refer to inhibition of expression of any complement factor Bgene (such as, e.g., a mouse complement factor B gene, a rat complementfactor B gene, a monkey complement factor B gene, or a human complementfactor B gene) as well as variants or mutants of a complement factor Bgene. Thus, the complement factor B gene may be a wild-type complementfactor B gene, a mutant complement factor B gene, or a transgeniccomplement factor B gene in the context of a genetically manipulatedcell, group of cells, or organism.

“Inhibiting expression of a complement factor B gene” includes any levelof inhibition of a complement factor B gene, e.g., at least partialsuppression of the expression of a complement factor B gene, such as aclinically relevant level of suppression. The expression of thecomplement factor B gene may be assessed based on the level, or thechange in the level, of any variable associated with complement factor Bgene expression, e.g., complement factor B mRNA level or complementfactor B protein level, or, for example, CH₅₀ activity as a measure oftotal hemolytic complement, AH₅₀ to measure the hemolytic activity ofthe alternate pathway of complement, or lactate dehydrogenase (LDH)levels as a measure of intravascular hemolysis, or hemoglobin levels.Levels of C3, C9, C5, C5a, C5b, and soluble C5b-9 complex may also bemeasured to assess CFB expression. Inhibition may be assessed by adecrease in an absolute or relative level of one or more of thesevariables compared with a control level. This level may be assessed inan individual cell or in a group of cells, including, for example, asample derived from a subject. It is understood that complement factor Bis expressed predominantly in the liver, and is present in circulation.

Inhibition may be assessed by a decrease in an absolute or relativelevel of one or more variables that are associated with complementfactor B expression compared with a control level. The control level maybe any type of control level that is utilized in the art, e.g., apre-dose baseline level, or a level determined from a similar subject,cell, or sample that is untreated or treated with a control (such as,e.g., buffer only control or inactive agent control).

In some embodiments of the methods of the invention, expression of acomplement factor B gene is inhibited by at least 50%, 55%, 60%, 65%,70%, 75%, 80%, 85%, 90%, or 95%, or to below the level of detection ofthe assay. In certain embodiments, expression of a complement factor Bgene is inhibited by at least 70%. It is further understood thatinhibition of complement factor B expression in certain tissues, e.g.,in gall bladder, without a significant inhibition of expression in othertissues, e.g., brain, may be desirable. In certain embodiments,expression level is determined using the assay method provided inExample 2 with a 10 nM siRNA concentration in the appropriate speciesmatched cell line.

In certain embodiments, inhibition of expression in vivo is determinedby knockdown of the human gene in a rodent expressing the human gene,e.g., an AAV-infected mouse expressing the human target gene (i.e.,complement factor B), e.g., when administered as a single dose, e.g., at3 mg/kg at the nadir of RNA expression. Knockdown of expression of anendogenous gene in a model animal system can also be determined, e.g.,after administration of a single dose at, e.g., 3 mg/kg at the nadir ofRNA expression. Such systems are useful when the nucleic acid sequenceof the human gene and the model animal gene are sufficiently close suchthat the human iRNA provides effective knockdown of the model animalgene. RNA expression in liver is determined using the PCR methodsprovided in Example 2.

Inhibition of the expression of a complement factor B gene may bemanifested by a reduction of the amount of mRNA expressed by a firstcell or group of cells (such cells may be present, for example, in asample derived from a subject) in which a complement factor B gene istranscribed and which has or have been treated (e.g., by contacting thecell or cells with an iRNA of the invention, or by administering an iRNAof the invention to a subject in which the cells are or were present)such that the expression of a complement factor B gene is inhibited, ascompared to a second cell or group of cells substantially identical tothe first cell or group of cells but which has not or have not been sotreated (control cell(s) not treated with an iRNA or not treated with aniRNA targeted to the gene of interest). In certain embodiments, theinhibition is assessed by the method provided in Example 2 using a 10 nMsiRNA concentration in the species matched cell line and expressing thelevel of mRNA in treated cells as a percentage of the level of mRNA incontrol cells, using the following formula:

${\frac{\left( {{mRNA}{in}{control}{cells}} \right) - \left( {{mRNA}{in}{treated}{cells}} \right)}{\left( {{mRNA}{in}{control}{cells}} \right)} \cdot 100}\%$

In other embodiments, inhibition of the expression of a complementfactor B gene may be assessed in terms of a reduction of a parameterthat is functionally linked to complement factor B gene expression,e.g., complement factor B protein level in blood or serum from asubject. Complement factor B gene silencing may be determined in anycell expressing complement factor B, either endogenous or heterologousfrom an expression construct, and by any assay known in the art.

Inhibition of the expression of a complement factor B protein may bemanifested by a reduction in the level of the complement factor Bprotein that is expressed by a cell or group of cells or in a subjectsample (e.g., the level of protein in a blood sample derived from asubject). As explained above, for the assessment of mRNA suppression,the inhibition of protein expression levels in a treated cell or groupof cells may similarly be expressed as a percentage of the level ofprotein in a control cell or group of cells, or the change in the levelof protein in a subject sample, e.g., blood or serum derived therefrom.

A control cell, a group of cells, or subject sample that may be used toassess the inhibition of the expression of a complement factor B geneincludes a cell, group of cells, or subject sample that has not yet beencontacted with an RNAi agent of the invention. For example, the controlcell, group of cells, or subject sample may be derived from anindividual subject (e.g., a human or animal subject) prior to treatmentof the subject with an RNAi agent or an appropriately matched populationcontrol.

The level of complement factor B mRNA that is expressed by a cell orgroup of cells may be determined using any method known in the art forassessing mRNA expression. In one embodiment, the level of expression ofcomplement factor B in a sample is determined by detecting a transcribedpolynucleotide, or portion thereof, e.g., mRNA of the complement factorB gene. RNA may be extracted from cells using RNA extraction techniquesincluding, for example, using acid phenol/guanidine isothiocyanateextraction (RNAzol B; Biogenesis), RNeasy™ RNA preparation kits(Qiagen®) or PAXgene™ (PreAnalytix™, Switzerland). Typical assay formatsutilizing ribonucleic acid hybridization include nuclear run-on assays,RT-PCR, RNase protection assays, northern blotting, in situhybridization, and microarray analysis.

In some embodiments, the level of expression of complement factor B isdetermined using a nucleic acid probe. The term “probe”, as used herein,refers to any molecule that is capable of selectively binding to aspecific complement factor B. Probes can be synthesized by one of skillin the art, or derived from appropriate biological preparations. Probesmay be specifically designed to be labeled. Examples of molecules thatcan be utilized as probes include, but are not limited to, RNA, DNA,proteins, antibodies, and organic molecules.

Isolated mRNA can be used in hybridization or amplification assays thatinclude, but are not limited to, Southern or northern analyses,polymerase chain reaction (PCR) analyses and probe arrays. One methodfor the determination of mRNA levels involves contacting the isolatedmRNA with a nucleic acid molecule (probe) that can hybridize tocomplement factor B mRNA. In one embodiment, the mRNA is immobilized ona solid surface and contacted with a probe, for example by running theisolated mRNA on an agarose gel and transferring the mRNA from the gelto a membrane, such as nitrocellulose. In an alternative embodiment, theprobe(s) are immobilized on a solid surface and the mRNA is contactedwith the probe(s), for example, in an Affymetrix® gene chip array. Askilled artisan can readily adapt known mRNA detection methods for usein determining the level of complement factor B mRNA.

An alternative method for determining the level of expression ofcomplement factor B in a sample involves the process of nucleic acidamplification or reverse transcriptase (to prepare cDNA) of for examplemRNA in the sample, e.g., by RT-PCR (the experimental embodiment setforth in Mullis, 1987, U.S. Pat. No. 4,683,202), ligase chain reaction(Barany (1991) Proc. Natl. Acad. Sci. USA 88:189-193), self sustainedsequence replication (Guatelli et al. (1990) Proc. Natl. Acad. Sci. USA87:1874-1878), transcriptional amplification system (Kwoh et al. (1989)Proc. Natl. Acad. Sci. USA 86:1173-1177), Q-Beta Replicase (Lizardi etal. (1988) Bio/Technology 6:1197), rolling circle replication (Lizardiet al., U.S. Pat. No. 5,854,033) or any other nucleic acid amplificationmethod, followed by the detection of the amplified molecules usingtechniques well known to those of skill in the art. These detectionschemes are especially useful for the detection of nucleic acidmolecules if such molecules are present in very low numbers. Inparticular aspects of the invention, the level of expression of CFB isdetermined by quantitative fluorogenic RT-PCR (i.e., the TaqMan™System). In certain embodiments, expression level is determined by themethod provided in Example 2 using, e.g., a 10 nM siRNA concentration,in the species matched cell line.

The expression levels of complement factor B mRNA may be monitored usinga membrane blot (such as used in hybridization analysis such asnorthern, Southern, dot, and the like), or microwells, sample tubes,gels, beads or fibers (or any solid support comprising bound nucleicacids). See U.S. Pat. Nos. 5,770,722, 5,874,219, 5,744,305, 5,677,195and 5,445,934, which are incorporated herein by reference. Thedetermination of complement factor B expression level may also compriseusing nucleic acid probes in solution.

In certain embodiments, the level of mRNA expression is assessed usingbranched DNA (bDNA) assays or real time PCR (qPCR). The use of thesemethods is described and exemplified in the Examples presented herein.In certain embodiments, expression level is determined by the methodprovided in Example 2 using a 10 nM siRNA concentration in the speciesmatched cell line.

The level of CFB protein expression may be determined using any methodknown in the art for the measurement of protein levels. Such methodsinclude, for example, electrophoresis, capillary electrophoresis, highperformance liquid chromatography (HPLC), thin layer chromatography(TLC), hyperdiffusion chromatography, fluid or gel precipitin reactions,absorption spectroscopy, a colorimetric assays, spectrophotometricassays, flow cytometry, immunodiffusion (single or double),immunoelectrophoresis, western blotting, radioimmunoassay (RIA),enzyme-linked immunosorbent assays (ELISAs), immunofluorescent assays,electrochemiluminescence assays, and the like.

In some embodiments, the efficacy of the methods of the invention isassessed by a decrease in CFB mRNA or protein level (e.g., in a liverbiopsy).

In some embodiments of the methods of the invention, the iRNA isadministered to a subject such that the iRNA is delivered to a specificsite within the subject. The inhibition of expression of complementfactor B may be assessed using measurements of the level or change inthe level of complement factor B mRNA or complement factor B protein ina sample derived from fluid or tissue from the specific site within thesubject (e.g., liver or blood).

As used herein, the terms detecting or determining a level of an analyteare understood to mean performing the steps to determine if a material,e.g., protein, RNA, is present. As used herein, methods of detecting ordetermining include detection or determination of an analyte level thatis below the level of detection for the method used.

VII. Prophylactic and Treatment Methods of the Invention

The present invention also provides methods of using an iRNA of theinvention or a composition containing an iRNA of the invention toinhibit expression of complement factor B, thereby preventing ortreating a complement factor B-associated disorder, e.g., paroxysmalnocturnal hemoglobinuria (PNH), atypical hemolytic uremic syndrome(aHUS), asthma, rheumatoid arthritis (RA); antiphospholipid antibodysyndrome; lupus nephritis; ischemia-reperfusion injury; typical orinfectious hemolytic uremic syndrome (tHUS); dense deposit disease(DDD); neuromyelitis optica (NMO); multifocal motor neuropathy (MMN);multiple sclerosis (MS); macular degeneration (e.g., age-related maculardegeneration (AMD)); hemolysis, elevated liver enzymes, and lowplatelets (HELLP) syndrome; thrombotic thrombocytopenic purpura (TTP);spontaneous fetal loss; Pauci-immune vasculitis; epidermolysis bullosa;recurrent fetal loss; pre-eclampsia, traumatic brain injury, myastheniagravis, cold agglutinin disease, dermatomyositis bullous pemphigoid,Shiga toxin E. coli-related hemolytic uremic syndrome, C3 neuropathy,anti-neutrophil cytoplasmic antibody-associated vasculitis (e.g.,granulomatosis with polyangiitis (previously known as Wegenergranulomatosis), Churg-Strauss syndrome, and microscopic polyangiitis),humoral and vascular transplant rejection, graft dysfunction, myocardialinfarction (e.g., tissue damage and ischemia in myocardial infarction),an allogenic transplant, sepsis (e.g., poor outcome in sepsis), Coronaryartery disease, dermatomyositis, Graves' disease, atherosclerosis,Alzheimer's disease, systemic inflammatory response sepsis, septicshock, spinal cord injury, glomerulonephritis, Hashimoto's thyroiditis,type I diabetes, psoriasis, pemphigus, autoimmune hemolytic anemia(AIHA), ITP, Goodpasture syndrome, Degos disease, antiphospholipidsyndrome (APS), catastrophic APS (CAPS), a cardiovascular disorder,myocarditis, a cerebrovascular disorder, a peripheral (e.g.,musculoskeletal) vascular disorder, a renovascular disorder, amesenteric/enteric vascular disorder, vasculitis, Henoch-Schönleinpurpura nephritis, systemic lupus erythematosus-associated vasculitis,vasculitis associated with rheumatoid arthritis, immune complexvasculitis, Takayasu's disease, dilated cardiomyopathy, diabeticangiopathy, Kawasaki's disease (arteritis), venous gas embolus (VGE),and restenosis following stent placement, rotational atherectomy, andpercutaneous transluminal coronary angioplasty (PTCA) (see, e.g., Holers(2008) Immunological Reviews 223:300-316; Holers and Thurman (2004)Molecular Immunology 41:147-152; U.S. Patent Publication No.20070172483).

In one embodiment, the complement factor B-associate disease is selectedfrom the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, polycystic kidney disease, membranous nephropathy,age-related macular degeneration, atypical hemolytic uremic syndrome,thrombotic microangiopathy, myasthenia gravis, ischemia and reperfusioninjury, paroxysmal nocturnal hemoglobinuria, and rheumatoid arthritis

In another embodiment, the complement factor B-associate disease isselected from the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

In the methods of the invention the cell may be contacted with the siRNAin vitro or in vivo, i.e., the cell may be within a subject.

A cell suitable for treatment using the methods of the invention may beany cell that expresses a complement factor B gene, e.g., a liver cell,a brain cell, a gall bladder cell, a heart cell, or a kidney cell. Inone embodiment, the cell is a liver cell. A cell suitable for use in themethods of the invention may be a mammalian cell, e.g., a primate cell(such as a human cell, including human cell in a chimeric non-humananimal, or a non-human primate cell, e.g., a monkey cell or a chimpanzeecell), or a non-primate cell. In certain embodiments, the cell is ahuman cell, e.g., a human liver cell. In the methods of the invention,complement factor B expression is inhibited in the cell by at least 50,55, 60, 65, 70, 75, 80, 85, 90, or 95, or to a level below the level ofdetection of the assay.

The in vivo methods of the invention may include administering to asubject a composition containing an iRNA, where the iRNA includes anucleotide sequence that is complementary to at least a part of an RNAtranscript of the complement factor B gene of the mammal to which theRNAi agent is to be administered. The composition can be administered byany means known in the art including, but not limited to oral,intraperitoneal, or parenteral routes, including intracranial (e.g.,intraventricular, intraparenchymal, and intrathecal), intravenous,intramuscular, subcutaneous, transdermal, airway (aerosol), nasal,rectal, and topical (including buccal and sublingual) administration. Incertain embodiments, the compositions are administered by intravenousinfusion or injection. In certain embodiments, the compositions areadministered by subcutaneous injection. In certain embodiments, thecompositions are administered by intramuscular injection.

In some embodiments, the administration is via a depot injection. Adepot injection may release the iRNA in a consistent way over aprolonged time period. Thus, a depot injection may reduce the frequencyof dosing needed to obtain a desired effect, e.g., a desired inhibitionof CFB, or a therapeutic or prophylactic effect. A depot injection mayalso provide more consistent serum concentrations. Depot injections mayinclude subcutaneous injections or intramuscular injections. In certainembodiments, the depot injection is a subcutaneous injection.

In some embodiments, the administration is via a pump. The pump may bean external pump or a surgically implanted pump. In certain embodiments,the pump is a subcutaneously implanted osmotic pump. In otherembodiments, the pump is an infusion pump. An infusion pump may be usedfor intravenous, subcutaneous, arterial, or epidural infusions. Incertain embodiments, the infusion pump is a subcutaneous infusion pump.In other embodiments, the pump is a surgically implanted pump thatdelivers the iRNA to the liver.

The mode of administration may be chosen based upon whether local orsystemic treatment is desired and based upon the area to be treated. Theroute and site of administration may be chosen to enhance targeting.

In one aspect, the present invention also provides methods forinhibiting the expression of a complement factor B gene in a mammal. Themethods include administering to the mammal a composition comprising adsRNA that targets a complement factor B gene in a cell of the mammaland maintaining the mammal for a time sufficient to obtain degradationof the mRNA transcript of the complement factor B gene, therebyinhibiting expression of the complement factor B gene in the cell.Reduction in gene expression can be assessed by any methods known in theart and by methods, e.g. qRT-PCR, described herein, e.g., in Example 2.Reduction in protein production can be assessed by any methods known itthe art, e.g. ELISA. In certain embodiments, a puncture liver biopsysample serves as the tissue material for monitoring the reduction in thecomplement factor B gene or protein expression. In other embodiments, ablood sample serves as the subject sample for monitoring the reductionin the complement factor B protein expression.

The present invention further provides methods of treatment in a subjectin need thereof, e.g., a subject diagnosed with a complement factorB-associated disorder, such as, C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

The present invention further provides methods of prophylaxis in asubject in need thereof. The treatment methods of the invention includeadministering an iRNA of the invention to a subject, e.g., a subjectthat would benefit from a reduction of complement factor B expression,in a prophylactically effective amount of an iRNA targeting a complementfactor B gene or a pharmaceutical composition comprising an iRNAtargeting a complement factor B gene.

In one embodiment, a complement factor B-associated disease is selectedfrom the group consisting of paroxysmal nocturnal hemoglobinuria (PNH),atypical hemolytic uremic syndrome (aHUS), asthma, rheumatoid arthritis(RA); antiphospholipid antibody syndrome; lupus nephritis;ischemia-reperfusion injury; typical or infectious hemolytic uremicsyndrome (tHUS); dense deposit disease (DDD); neuromyelitis optica(NMO); multifocal motor neuropathy (MMN); multiple sclerosis (MS);macular degeneration (e.g., age-related macular degeneration (AMD));hemolysis, elevated liver enzymes, and low platelets (HELLP) syndrome;thrombotic thrombocytopenic purpura (TTP); spontaneous fetal loss;Pauci-immune vasculitis; epidermolysis bullosa; recurrent fetal loss;pre-eclampsia, traumatic brain injury, myasthenia gravis, coldagglutinin disease, dermatomyositis bullous pemphigoid, Shiga toxin E.coli-related hemolytic uremic syndrome, C3 neuropathy, anti-neutrophilcytoplasmic antibody-associated vasculitis (e.g., granulomatosis withpolyangiitis (previously known as Wegener granulomatosis), Churg-Strausssyndrome, and microscopic polyangiitis), humoral and vascular transplantrejection, graft dysfunction, myocardial infarction (e.g., tissue damageand ischemia in myocardial infarction), an allogenic transplant, sepsis(e.g., poor outcome in sepsis), Coronary artery disease,dermatomyositis, Graves' disease, atherosclerosis, Alzheimer's disease,systemic inflammatory response sepsis, septic shock, spinal cord injury,glomerulonephritis, Hashimoto's thyroiditis, type I diabetes, psoriasis,pemphigus, autoimmune hemolytic anemia (AIHA), ITP, Goodpasturesyndrome, Degos disease, antiphospholipid syndrome (APS), catastrophicAPS (CAPS), a cardiovascular disorder, myocarditis, a cerebrovasculardisorder, a peripheral (e.g., musculoskeletal) vascular disorder, arenovascular disorder, a mesenteric/enteric vascular disorder,vasculitis, Henoch-Schönlein purpura nephritis, systemic lupuserythematosus-associated vasculitis, vasculitis associated withrheumatoid arthritis, immune complex vasculitis, Takayasu's disease,dilated cardiomyopathy, diabetic angiopathy, Kawasaki's disease(arteritis), venous gas embolus (VGE), and restenosis following stentplacement, rotational atherectomy, and percutaneous transluminalcoronary angioplasty (PTCA) (see, e.g., Holers (2008) ImmunologicalReviews 223:300-316; Holers and Thurman (2004) Molecular Immunology41:147-152; US20070172483).

In one embodiment, the complement factor B-associate disease is selectedfrom the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, polycystic kidney disease, membranous nephropathy,age-related macular degeneration, atypical hemolytic uremic syndrome,thrombotic microangiopathy, myasthenia gravis, ischemia and reperfusioninjury, paroxysmal nocturnal hemoglobinuria, and rheumatoid arthritis

In another embodiment, the complement factor B-associate disease isselected from the group consisting of C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

An iRNA of the invention may be administered as a “free iRNA.” A freeiRNA is administered in the absence of a pharmaceutical composition. Thenaked iRNA may be in a suitable buffer solution. The buffer solution maycomprise acetate, citrate, prolamine, carbonate, or phosphate, or anycombination thereof. In one embodiment, the buffer solution is phosphatebuffered saline (PBS). The pH and osmolarity of the buffer solutioncontaining the iRNA can be adjusted such that it is suitable foradministering to a subject.

Alternatively, an iRNA of the invention may be administered as apharmaceutical composition, such as a dsRNA liposomal formulation.

Subjects that would benefit from an inhibition of complement factor Bgene expression are subjects susceptible to or diagnosed with aCFB-associated disorder, e.g., C3 glomerulopathy, systemic lupuserythematosus (SLE), e.g., Lupus Nephritis, IgA nephropathy, diabeticnephropathy, and polycystic kidney disease.

In an embodiment, the method includes administering a compositionfeatured herein such that expression of the target complement componentB gene is decreased, such as for about 1, 2, 3, 4, 5, 6, 1-6, 1-3, or3-6 months per dose. In certain embodiments, the composition isadministered once every 3-6 months.

In one embodiment, the iRNAs useful for the methods and compositionsfeatured herein specifically target RNAs (primary or processed) of thetarget complement factor B gene. Compositions and methods for inhibitingthe expression of these genes using iRNAs can be prepared and performedas described herein.

Administration of the iRNA according to the methods of the invention mayresult prevention or treatment of a complement factor B-associateddisorder, e.g., C3 glomerulopathy, systemic lupus erythematosus (SLE),e.g., Lupus Nephritis, IgA nephropathy, diabetic nephropathy, andpolycystic kidney disease.

Subjects can be administered a therapeutic amount of iRNA, such as about0.01 mg/kg to about 200 mg/kg. Subjects can be administered atherapeutic amount of iRNA, such as about 5 mg to about 1000 mg as afixed dose, regardless of body weight.

In some embodiment, the iRNA is administered subcutaneously, i.e., bysubcutaneous injection. One or more injections may be used to deliverthe desired dose of iRNA to a subject. The injections may be repeatedover a period of time.

The administration may be repeated on a regular basis. In certainembodiments, after an initial treatment regimen, the treatments can beadministered on a less frequent basis. A repeat-dose regimen may includeadministration of a therapeutic amount of iRNA on a regular basis, suchas once per month to once a year. In certain embodiments, the iRNA isadministered about once per month to about once every three months, orabout once every three months to about once every six months.

The invention further provides methods and uses of an iRNA agent or apharmaceutical composition thereof for treating a subject that wouldbenefit from reduction or inhibition of CFB gene expression, e.g., asubject having a CFB-associated disease, in combination with otherpharmaceuticals or other therapeutic methods, e.g., with knownpharmaceuticals or known therapeutic methods, such as, for example,those which are currently employed for treating these disorders.

Accordingly, in some aspects of the invention, the methods which includeeither a single iRNA agent of the invention, further includeadministering to the subject one or more additional therapeutic agents.The iRNA agent and an additional therapeutic agent or treatment may beadministered at the same time or in the same combination, e.g.,parenterally, or the additional therapeutic agent can be administered aspart of a separate composition or at separate times or by another methodknown in the art or described herein.

In one embodiment, an iRNA agent of the invention is administered incombination with an anti-complement component C5 antibody, orantigen-binding fragment thereof (e.g., eculizumab or ravulizumab-cwvz),an iRNA agent targeting complement component C5, an iRNA agent targetingcomplement component C3, or a C3 peptide inhibitor (e.g., compstatin).In one embodiment, the iRNA agent of the invention is administered tothe patient, and then the additional therapeutic agent is administeredto the patient (or vice versa). In another embodiment, the iRNA agent ofthe invention and the additional therapeutic agent are administered atthe same time.

The iRNA agent of the invention and an additional therapeutic agent ortreatment may be administered at the same time or in the samecombination, e.g., parenterally, or the additional therapeutic agent canbe administered as part of a separate composition or at separate timesor by another method known in the art or described herein.

VIII. Kits

In certain aspects, the instant disclosure provides kits that include asuitable container containing a pharmaceutical formulation of a siRNAcompound, e.g., a double-stranded siRNA compound, or ssiRNA compound,(e.g., a precursor, e.g., a larger siRNA compound which can be processedinto a ssiRNA compound, or a DNA which encodes an siRNA compound, e.g.,a double-stranded siRNA compound, or ssiRNA compound, or precursorthereof).

Such kits include one or more dsRNA agent(s) and instructions for use,e.g., instructions for administering a prophylactically ortherapeutically effective amount of a dsRNA agent(s). The dsRNA agentmay be in a vial or a pre-filled syringe. The kits may optionallyfurther comprise means for administering the dsRNA agent (e.g., aninjection device, such as a pre-filled syringe), or means for measuringthe inhibition of CFB (e.g., means for measuring the inhibition of CFBmRNA, CFB protein, or CFB activity). Such means for measuring theinhibition of CFB may comprise a means for obtaining a sample from asubject, such as, e.g., a plasma sample. The kits of the invention mayoptionally further comprise means for determining the therapeuticallyeffective or prophylactically effective amount.

In certain embodiments the individual components of the pharmaceuticalformulation may be provided in one container, e.g., a vial or apre-filled syringe. Alternatively, it may be desirable to provide thecomponents of the pharmaceutical formulation separately in two or morecontainers, e.g., one container for a siRNA compound preparation, and atleast another for a carrier compound. The kit may be packaged in anumber of different configurations such as one or more containers in asingle box. The different components can be combined, e.g., according toinstructions provided with the kit. The components can be combinedaccording to a method described herein, e.g., to prepare and administera pharmaceutical composition. The kit can also include a deliverydevice.

This invention is further illustrated by the following examples whichshould not be construed as limiting. The entire contents of allpublications, patents and published patent applications cited throughoutthis application, as well as the informal Sequence Listing and Figures,are hereby incorporated herein by reference.

EXAMPLES Example 1. iRNA Synthesis Source of Reagents

Where the source of a reagent is not specifically given herein, suchreagent can be obtained from any supplier of reagents for molecularbiology at a quality/purity standard for application in molecularbiology.

siRNA Design

siRNAs targeting the human complement factor B (CFB) gene, (human: NCBIrefseqID NM_001710.6; NCBI GeneID: 629) was designed using custom RandPython scripts. The human NM_001710 REFSEQ mRNA, version 6, has a lengthof 2476 bases. Detailed lists of the unmodified CFB sense and antisensestrand nucleotide sequences are shown in Table 2. Detailed lists of themodified CFB sense and antisense strand nucleotide sequences are shownin Table 3.

It is to be understood that, throughout the application, a duplex namewithout a decimal is equivalent to a duplex name with a decimal whichmerely references the batch number of the duplex. For example, AD-959917is equivalent to AD-959917.1.

siRNA Synthesis

siRNAs were synthesized and annealed using routine methods known in theart.

Briefly, siRNA sequences were synthesized on a 1 μmol scale using aMermade 192 synthesizer (BioAutomation) with phosphoramidite chemistryon solid supports. The solid support was controlled pore glass (500-1000Å) loaded with a custom GalNAc ligand (3′-GalNAc conjugates), universalsolid support (AM Chemicals), or the first nucleotide of interest.Ancillary synthesis reagents and standard 2-cyanoethyl phosphoramiditemonomers (2′-deoxy-2′-fluoro, 2′-O-methyl, RNA, DNA) were obtained fromThermo-Fisher (Milwaukee, Wis.), Hongene (China), or Chemgenes(Wilmington, Mass., USA). Additional phosphoramidite monomers wereprocured from commercial suppliers, prepared in-house, or procured usingcustom synthesis from various CMOs. Phosphoramidites were prepared at aconcentration of 100 mM in either acetonitrile or 9:1 acetonitrile:DMFand were coupled using 5-Ethylthio-1H-tetrazole (ETT, 0.25 M inacetonitrile) with a reaction time of 400 s. Phosphorothioate linkageswere generated using a 100 mM solution of 3-((Dimethylamino-methylidene)amino)-3H-1,2,4-dithiazole-3-thione (DDTT, obtained from Chemgenes(Wilmington, Mass., USA)) in anhydrous acetonitrile/pyridine (9:1 v/v).Oxidation time was 5 minutes. All sequences were synthesized with finalremoval of the DMT group (“DMT-Off”).

Upon completion of the solid phase synthesis, solid-supportedoligoribonucleotides were treated with 300 μL of Methylamine (40%aqueous) at room temperature in 96 well plates for approximately 2 hoursto afford cleavage from the solid support and subsequent removal of alladditional base-labile protecting groups. For sequences containing anynatural ribonucleotide linkages (2′-OH) protected with a tert-butyldimethyl silyl (TBDMS) group, a second deprotection step was performedusing TEA·3HF (triethylamine trihydrofluoride). To each oligonucleotidesolution in aqueous methylamine was added 200 μL of dimethyl sulfoxide(DMSO) and 300 μL TEA·3HF and the solution was incubated forapproximately 30 mins at 60° C. After incubation, the plate was allowedto come to room temperature and crude oligonucleotides were precipitatedby the addition of 1 mL of 9:1 acetontrile:ethanol or 1:1ethanol:isopropanol. The plates were then centrifuged at 4° C. for 45mins and the supernatant carefully decanted with the aid of amultichannel pipette. The oligonucleotide pellet was resuspended in 20mM NaOAc and subsequently desalted using a HiTrap size exclusion column(5 mL, GE Healthcare) on an Agilent LC system equipped with anautosampler, UV detector, conductivity meter, and fraction collector.Desalted samples were collected in 96 well plates and then analyzed byLC-MS and UV spectrometry to confirm identity and quantify the amount ofmaterial, respectively.

Duplexing of single strands was performed on a Tecan liquid handlingrobot. Sense and antisense single strands were combined in an equimolarratio to a final concentration of 10 μM in 1×PBS in 96 well plates, theplate sealed, incubated at 100° C. for 10 minutes, and subsequentlyallowed to return slowly to room temperature over a period of 2-3 hours.The concentration and identity of each duplex was confirmed and thensubsequently utilized for in vitro screening assays.

TABLE 1 Abbreviations of nucleotide monomers used in nucleic acidsequence representation. It will be understood that these monomers, whenpresent in an oligonucleotide, are mutually linked by 5′-3′-phosphodiester bonds; and it is understood that when the nucleotidecontains a 2′-fluoro modification, then the fluoro replaces the hydroxyat that position in the parent nucleotide (i.e., it is a 2′-deoxy-2′-fluoronucleotide). It is to be further understood that the nucleotideabbreviations in the table omit the 3′-phosphate (i.e., they are 3′-OH)when placed at the 3′-terminal position of an oligonucleotide.Abbreviation Nucleotide(s) A Adenosine-3′-phosphate Abbeta-L-adenosine-3′-phosphate Abs beta-L-adenosine-3′-phosphorothioateAf 2′-fluoroadenosine-3'-phosphate Afs2′-fluoroadenosine-3′-phosphorothioate As adenosine-3′-phosphorothioateC cytidine-3'-phosphate Cb beta-L-cytidine-3′-phosphate Cbsbeta-L-cytidine-3′-phosphorothioate Cf 2′-fluorocytidine-3′-phosphateCfs 2′-fluorocytidine-3′-phosphorothioate Cscytidine-3′-phosphorothioate G guanosine-3′-phosphate Gbbeta-L-guanosine-3′-phosphate Gbs beta-L-guanosine-3′-phosphorothioateGf 2′-fluoroguanosine-3′-phosphate Gfs2′-fluoroguanosine-3′-phosphorothioate Gs guanosine-3′-phosphorothioateT 5′-methyluridine-3′-phosphate Tf2′-fluoro-5-methyluridine-3′-phosphate Tfs2′-fluoro-5-methyluridine-3′-phosphorothioate Ts5-methyluridine-3′-phosphorothioate U Uridine-3′-phosphate Uf2′-fluorouridine-3′-phosphate Ufs 2′-fluorouridine-3′-phosphorothioateUs uridine-3′-phosphorothioate N any nucleotide, modified or unmodifieda 2′-O-methyladenosine-3′-phosphate as2′-O-methyladenosine-3′-phosphorothioate c2′-O-methylcytidine-3′-phosphate cs2′-O-methylcytidine-3′-phosphorothioate g2′-O-methylguanosine-3′-phosphate gs2′-O-methylguanosine-3′-phosphorothioate t2′-O-methyl-5-methyluridine-3′-phosphate ts2′-O-methyl-5-methyluridine-3′-phosphorothioate u2′-O-methyluridine-3′-phosphate us2′-O-methyluridine-3′-phosphorothioate S phosphorothioate linkage L96N-[tris(GalNAc-alkyl)-amidodecanoyl)]-4-hydroxyprolinol(Hyp-(GalNAc-alkyl)3)(2S,4R)-1-[29-[[2-(acetylamino)-2-deoxy-ß-D-galactopyranosyl]oxy]-14,14-bis[[3-[[3-[[5-[[2-(acetylamino)-2-deoxy-ß-D-galactopyranosyl]oxy]-1-oxopentyl]amino]propyl]amino]-3-oxopropoxy]methyl]-1,12,19,25-tetraoxo-16-oxa-13,20,24-triazanonacos-1-yl]-4-hydroxy-2-hydroxymethylpyrrolidine

uL96 2′-O-methyluridine-3'-phosphate ((2S,4R)-1-[29-[[2-(acetylamino)-2-deoxy-ß-D-galactopyranosyl]oxy]-14,14-bis[[3-[[3-[[5-[[2-(acetylamino)-2-deoxy-ß-D-galactopyranosyl]oxy]-1-oxopentyl]amino]propyl]amino]-3-oxopropoxy]methyl]-1,12,19,25-tetraoxo-16-oxa-13,20,24-triazanonacos-1-yl]-4-hydroxy-2-pyrrolidinyl)methyl ester

Y34 2-hydroxymethyl-tetrahydrofurane-4-methoxy-3-phosphate (abasic2′-OMe furanose)

Y44 inverted abasic DNA (2-hydroxymethyl-tetrahydrofurane-5-phosphate)

L10 N-(cholesterylcarboxamidocaproyl)-4-hydroxyprolinol (Hyp-C6-Chol)

(Agn) Adenosine-glycol nucleic acid (GNA) S-Isomer (Cgn) Cytidine-glycolnucleic acid (GNA) S-Isomer (Ggn) Guanosine-glycol nucleic acid (GNA)S-Isomer (Tgn) Thymidine-glycol nucleic acid (GNA) S-Isomer P PhosphateVP Vinyl-phosphonate dA 2′-deoxyadenosine-3′-phosphate dAs2′-deoxyadenosine-3′-phosphorothioate dC 2′-deoxycytidine-3′-phosphatedCs 2`-deoxycytidine-3`-phosphorothioate dG2′-deoxyguanosine-3′-phosphate dGs 2′-deoxyguanosine-3′-phosphorothioatedT 2′-deoxythymidine-3′-phosphate dTs2′-deoxythymidine-3′-phosphorothioate dU 2′-deoxyuridine dUs2′-deoxyuridine-3′-phosphorothioate (C2p) cytidine-2′-phosphate (G2p)guanosine-2′-phosphate (U2p) uridine-2′-phosphate (A2p)adenosine-2′-phosphate (Ahd) 2′-O-hexadecyl-adenosine-3′-phosphate(Ahds) 2′-O-hexadecyl-adenosine-3′-phosphorothioate (Chd)2′-O-hexadecyl-cytidine-3′-phosphate (Chds)2′-O-hexadecyl-cytidine-3′-phosphorothioate (Ghd)2′-O-hexadecyl-guanosine-3′-phosphate (Ghds)2′-O-hexadecyl-guanosine-3′-phosphorothioate (Uhd)2′-O-hexadecyl-uridine-3′-phosphate (Uhds)2′-O-hexadecyl-uridine-3′-phosphorothioate

TABLE 2Unmodified Sense and Antisense Strand Sequences of Complement Factor B dsRNA AgentsSEQ SEQ Duplex Sense Strand Sequence  ID Range inAntisense Strand Sequence ID Range in Name 5′ to 3′ NO: NM_001710.65′ to 3′ NO: NM_001710.6 AD-1724362 AAGGGAAUGUGACCAGGUCUU 19    4-24AAGACCTGGUCACAUUCCCUUCC 412    2-24 AD-1724363 AGGGAAUGUGACCAGGUCUAU 20   5-25 ATAGACCUGGUCACAUUCCCUUC 413    3-25 AD-1724364GGGAAUGUGACCAGGUCUAGU 21    6-26 ACUAGACCUGGTCACAUUCCCUU 414    4-26AD-1724365 GGAAUGUGACCAGGUCUAGGU 22    7-27 ACCUAGACCUGGUCACAUUCCCU 415   5-27 AD-1724369 UGUGACCAGGUCUAGGUCUGU 23   11-31ACAGACCUAGACCUGGUCACAUU 416    9-31 AD-1724370 GUGACCAGGUCUAGGUCUGGU 24  12-32 ACCAGACCUAGACCUGGUCACAU 417   10-32 AD-1724376AGGUCUAGGUCUGGAGUUUCU 25   18-38 AGAAACTCCAGACCUAGACCUGG 418   16-38AD-1724384 GUCUGGAGUUUCAGCUUGGAU 26   26-46 ATCCAAGCUGAAACUCCAGACCU 419  24-46 AD-1724385 UCUGGAGUUUCAGCUUGGACU 27   27-47AGUCCAAGCUGAAACUCCAGACC 420   25-47 AD-1724386 CUGGAGUUUCAGCUUGGACAU 28  28-48 ATGUCCAAGCUGAAACUCCAGAC 421   26-48 AD-1724530UCCUUCCGACUUCUCCAAGAU 29  269-289 ATCUTGGAGAAGUCGGAAGGAGC 422  267-289AD-1724572 UGUCCUUCUGGCUUCUACCCU 30  311-331 AGGGTAGAAGCCAGAAGGACACA 423 309-331 AD-1724574 UCCUUCUGGCUUCUACCCGUU 31  313-333AACGGGTAGAAGCCAGAAGGACA 424  311-333 AD-1724575 CCUUCUGGCUUCUACCCGUAU 32 314-334 ATACGGGUAGAAGCCAGAAGGAC 425  312-334 AD-1724576CUUCUGGCUUCUACCCGUACU 33  315-335 AGUACGGGUAGAAGCCAGAAGGA 426  313-335AD-1724579 CUGGCUUCUACCCGUACCCUU 34  318-338 AAGGGUACGGGTAGAAGCCAGAA 427 316-338 AD-1724586 CUACCCGUACCCUGUGCAGAU 35  325-345ATCUGCACAGGGUACGGGUAGAA 428  323-345 AD-1724600 UGCAGACACGUACCUGCAGAU 36 339-359 ATCUGCAGGUACGUGUCUGCACA 429  337-359 AD-1724651AAGGCAGAGUGCAGAGCAAUU 37  410-430 AAUUGCTCUGCACUCUGCCUUCC 430  408-430AD-1724653 GGCAGAGUGCAGAGCAAUCCU 38  412-432 AGGATUGCUCUGCACUCUGCCUU 431 410-432 AD-1724685 CGGUCUCCCUACUACAAUGUU 39  476-496AACATUGUAGUAGGGAGACCGGG 432  474-496 AD-1724691 CCCUACUACAAUGUGAGUGAU 40 482-502 ATCACUCACAUTGUAGUAGGGAG 433  480-502 AD-1724692CCUACUACAAUGUGAGUGAUU 41  483-503 AAUCACTCACATUGUAGUAGGGA 434  481-503AD-1724693 CUACUACAAUGUGAGUGAUGU 42  484-504 ACAUCACUCACAUUGUAGUAGGG 435 482-504 AD-1724695 ACUACAAUGUGAGUGAUGAGU 43  486-506ACUCAUCACUCACAUUGUAGUAG 436  484-506 AD-1724698 ACAAUGUGAGUGAUGAGAUCU 44 489-509 AGAUCUCAUCACUCACAUUGUAG 437  487-509 AD-1724699CAAUGUGAGUGAUGAGAUCUU 45  490-510 AAGATCTCAUCACUCACAUUGUA 438  488-510AD-1724700 AAUGUGAGUGAUGAGAUCUCU 46  491-511 AGAGAUCUCAUCACUCACAUUGU 439 489-511 AD-1724701 AUGUGAGUGAUGAGAUCUCUU 47  492-512AAGAGATCUCATCACUCACAUUG 440  490-512 AD-1724702 UGUGAGUGAUGAGAUCUCUUU 48 493-513 AAAGAGAUCUCAUCACUCACAUU 441  491-513 AD-1724703GUGAGUGAUGAGAUCUCUUUU 49  494-514 AAAAGAGAUCUCAUCACUCACAU 442  492-514AD-1724704 UGAGUGAUGAGAUCUCUUUCU 50  495-515 AGAAAGAGAUCTCAUCACUCACA 443 493-515 AD-1724705 GAGUGAUGAGAUCUCUUUCCU 51  496-516AGGAAAGAGAUCUCAUCACUCAC 444  494-516 AD-1724706 AGUGAUGAGAUCUCUUUCCAU 52 497-517 ATGGAAAGAGATCUCAUCACUCA 445  495-517 AD-1724707GUGAUGAGAUCUCUUUCCACU 53  498-518 AGUGGAAAGAGAUCUCAUCACUC 446  496-518AD-1724708 UGAUGAGAUCUCUUUCCACUU 54  499-519 AAGUGGAAAGAGAUCUCAUCACU 447 497-519 AD-1724714 GAUCUCUUUCCACUGCUAUGU 55  505-525ACAUAGCAGUGGAAAGAGAUCUC 448  503-525 AD-1724715 AUCUCUUUCCACUGCUAUGAU 56 506-526 ATCATAGCAGUGGAAAGAGAUCU 449  504-526 AD-1724716UCUCUUUCCACUGCUAUGACU 57  507-527 AGUCAUAGCAGTGGAAAGAGAUC 450  505-527AD-1724717 CUCUUUCCACUGCUAUGACGU 58  508-528 ACGUCATAGCAGUGGAAAGAGAU 451 506-528 AD-1724718 UCUUUCCACUGCUAUGACGGU 59  509-529ACCGTCAUAGCAGUGGAAAGAGA 452  507-529 AD-1724725 ACUGCUAUGACGGUUACACUU 60 516-536 AAGUGUAACCGTCAUAGCAGUGG 453  514-536 AD-1724726CUGCUAUGACGGUUACACUCU 61  517-537 AGAGTGTAACCGUCAUAGCAGUG 454  515-537AD-1724730 UAUGACGGUUACACUCUCCGU 62  521-541 ACGGAGAGUGUAACCGUCAUAGC 455 519-541 AD-1724731 AUGACGGUUACACUCUCCGGU 63  522-542ACCGGAGAGUGTAACCGUCAUAG 456  520-542 AD-1724741 AUCGCACCUGCCAAGUGAAUU 64 552-572 AAUUCACUUGGCAGGUGCGAUUG 457  550-572 AD-1724742UCGCACCUGCCAAGUGAAUGU 65  553-573 ACAUTCACUUGGCAGGUGCGAUU 458  551-573AD-1724743 CGCACCUGCCAAGUGAAUGGU 66  554-574 ACCATUCACUUGGCAGGUGCGAU 459 552-574 AD-1724776 CAGACAGCGAUCUGUGACAAU 67  587-607ATUGTCACAGATCGCUGUCUGCC 460  585-607 AD-1724777 AGACAGCGAUCUGUGACAACU 68 588-608 AGUUGUCACAGAUCGCUGUCUGC 461  586-608 AD-1724778GACAGCGAUCUGUGACAACGU 69  589-609 ACGUTGTCACAGAUCGCUGUCUG 462  587-609AD-1724779 ACAGCGAUCUGUGACAACGGU 70  590-610 ACCGTUGUCACAGAUCGCUGUCU 463 588-610 AD-1724780 CAGCGAUCUGUGACAACGGAU 71  591-611ATCCGUTGUCACAGAUCGCUGUC 464  589-611 AD-1724781 AGCGAUCUGUGACAACGGAGU 72 592-612 ACUCCGTUGUCACAGAUCGCUGU 465  590-612 AD-1724792UGGCACAAGGAAGGUGGGCAU 73  643-663 ATGCCCACCUUCCUUGUGCCAAU 466  641-663AD-1724819 CCGCCUUGAAGACAGCGUCAU 74  670-690 ATGACGCUGUCTUCAAGGCGGUA 467 668-690 AD-1724823 CUUGAAGACAGCGUCACCUAU 75  674-694ATAGGUGACGCTGUCUUCAAGGC 468  672-694 AD-1724824 UUGAAGACAGCGUCACCUACU 76 675-695 AGUAGGTGACGCUGUCUUCAAGG 469  673-695 AD-1724825UGAAGACAGCGUCACCUACCU 77  676-696 AGGUAGGUGACGCUGUCUUCAAG 470  674-696AD-1724860 GUGUCAGGAAGGUGGCUCUUU 78  739-759 AAAGAGCCACCTUCCUGACACGU 471 737-759 AD-1724894 CCUUCCUGCCAAGACUCCUUU 79  773-793AAAGGAGUCUUGGCAGGAAGGCU 472  771-793 AD-1724897 UCCUGCCAAGACUCCUUCAUU 80 776-796 AAUGAAGGAGUCUUGGCAGGAAG 473  774-796 AD-1724899CUGCCAAGACUCCUUCAUGUU 81  778-798 AACATGAAGGAGUCUUGGCAGGA 474  776-798AD-1724900 UGCCAAGACUCCUUCAUGUAU 82  779-799 ATACAUGAAGGAGUCUUGGCAGG 475 777-799 AD-1724903 CAAGACUCCUUCAUGUACGAU 83  782-802ATCGTACAUGAAGGAGUCUUGGC 476  780-802 AD-1724904 AAGACUCCUUCAUGUACGACU 84 783-803 AGUCGUACAUGAAGGAGUCUUGG 477  781-803 AD-1724905AGACUCCUUCAUGUACGACAU 85  784-804 ATGUCGTACAUGAAGGAGUCUUG 478  782-804AD-1724906 GACUCCUUCAUGUACGACACU 86  785-805 AGUGTCGUACATGAAGGAGUCUU 479 783-805 AD-1724910 CAAGAGGUGGCCGAAGCUUUU 87  809-829AAAAGCTUCGGCCACCUCUUGAG 480  807-829 AD-1724919 GCCGAAGCUUUCCUGUCUUCU 88 818-838 AGAAGACAGGAAAGCUUCGGCCA 481  816-838 AD-1724945AGAGACCAUAGAAGGAGUCGU 89  844-864 ACGACUCCUUCTAUGGUCUCUGU 482  842-864AD-1724946 GAGACCAUAGAAGGAGUCGAU 90  845-865 ATCGACTCCUUCUAUGGUCUCUG 483 843-865 AD-1724947 AGACCAUAGAAGGAGUCGAUU 91  846-866AAUCGACUCCUTCUAUGGUCUCU 484  844-866 AD-1724948 GACCAUAGAAGGAGUCGAUGU 92 847-867 ACAUCGACUCCTUCUAUGGUCUC 485  845-867 AD-1724949ACCAUAGAAGGAGUCGAUGCU 93  848-868 AGCATCGACUCCUUCUAUGGUCU 486  846-868AD-1725000 CCUUCAGGCUCCAUGAACAUU 94  920-940 AAUGTUCAUGGAGCCUGAAGGGU 487 918-940 AD-1725003 UCAGGCUCCAUGAACAUCUAU 95  923-943ATAGAUGUUCATGGAGCCUGAAG 488  921-943 AD-1725004 CAGGCUCCAUGAACAUCUACU 96 924-944 AGUAGATGUUCAUGGAGCCUGAA 489  922-944 AD-1725013UGAACAUCUACCUGGUGCUAU 97  933-953 ATAGCACCAGGTAGAUGUUCAUG 490  931-953AD-1725015 AACAUCUACCUGGUGCUAGAU 98  935-955 ATCUAGCACCAGGUAGAUGUUCA 491 933-955 AD-1725017 CAUCUACCUGGUGCUAGAUGU 99  937-957ACAUCUAGCACCAGGUAGAUGUU 492  935-957 AD-1725018 AUCUACCUGGUGCUAGAUGGU100  938-958 ACCATCTAGCACCAGGUAGAUGU 493  936-958 AD-1725019UCUACCUGGUGCUAGAUGGAU 101  939-959 ATCCAUCUAGCACCAGGUAGAUG 494  937-959AD-1725020 CUACCUGGUGCUAGAUGGAUU 102  940-960 AAUCCATCUAGCACCAGGUAGAU495  938-960 AD-1725021 UACCUGGUGCUAGAUGGAUCU 103  941-961AGAUCCAUCUAGCACCAGGUAGA 496  939-961 AD-1725022 ACCUGGUGCUAGAUGGAUCAU104  942-962 ATGATCCAUCUAGCACCAGGUAG 497  940-962 AD-1725023CCUGGUGCUAGAUGGAUCAGU 105  943-963 ACUGAUCCAUCTAGCACCAGGUA 498  941-963AD-1725025 UGGUGCUAGAUGGAUCAGACU 106  945-965 AGUCTGAUCCATCUAGCACCAGG499  943-965 AD-1725027 GUGCUAGAUGGAUCAGACAGU 107  947-967ACUGTCTGAUCCAUCUAGCACCA 500  945-967 AD-1725028 UGCUAGAUGGAUCAGACAGCU108  948-968 AGCUGUCUGAUCCAUCUAGCACC 501  946-968 AD-1725033GAUGGAUCAGACAGCAUUGGU 109  953-973 ACCAAUGCUGUCUGAUCCAUCUA 502  951-973AD-1725039 CAACUUCACAGGAGCCAAAAU 110  979-999 ATUUTGGCUCCTGUGAAGUUGCU503  977-999 AD-1725040 AACUUCACAGGAGCCAAAAAU 111  980-1000ATUUTUGGCUCCUGUGAAGUUGC 504  978-1000 AD-1725041 ACUUCACAGGAGCCAAAAAGU112  981-1001 ACUUTUTGGCUCCUGUGAAGUUG 505  979-1001 AD-1725042CUUCACAGGAGCCAAAAAGUU 113  982-1002 AACUTUTUGGCTCCUGUGAAGUU 506 980-1002 AD-1725043 UUCACAGGAGCCAAAAAGUGU 114  983-1003ACACTUTUUGGCUCCUGUGAAGU 507  981-1003 AD-1725044 UCACAGGAGCCAAAAAGUGUU115  984-1004 AACACUTUUUGGCUCCUGUGAAG 508  982-1004 AD-1725045CACAGGAGCCAAAAAGUGUCU 116  985-1005 AGACACTUUUUGGCUCCUGUGAA 509 983-1005 AD-1725046 ACAGGAGCCAAAAAGUGUCUU 117  986-1006AAGACACUUUUTGGCUCCUGUGA 510  984-1006 AD-1725047 CAGGAGCCAAAAAGUGUCUAU118  987-1007 ATAGACACUUUTUGGCUCCUGUG 511  985-1007 AD-1725048AGGAGCCAAAAAGUGUCUAGU 119  988-1008 ACUAGACACUUTUUGGCUCCUGU 512 986-1008 AD-1725049 GGAGCCAAAAAGUGUCUAGUU 120  989-1009AACUAGACACUTUUUGGCUCCUG 513  987-1009 AD-1725050 GAGCCAAAAAGUGUCUAGUCU121  990-1010 AGACTAGACACTUUUUGGCUCCU 514  988-1010 AD-1725051AGCCAAAAAGUGUCUAGUCAU 122  991-1011 ATGACUAGACACUUUUUGGCUCC 515 989-1011 AD-1725052 GCCAAAAAGUGUCUAGUCAAU 123  992-1012ATUGACTAGACACUUUUUGGCUC 516  990-1012 AD-1725053 CCAAAAAGUGUCUAGUCAACU124  993-1013 AGUUGACUAGACACUUUUUGGCU 517  991-1013 AD-1725054CAAAAAGUGUCUAGUCAACUU 125  994-1014 AAGUTGACUAGACACUUUUUGGC 518 992-1014 AD-1725055 AAAAAGUGUCUAGUCAACUUU 126  995-1015AAAGTUGACUAGACACUUUUUGG 519  993-1015 AD-1725056 AAAAGUGUCUAGUCAACUUAU127  996-1016 ATAAGUTGACUAGACACUUUUUG 520  994-1016 AD-1725057AAAGUGUCUAGUCAACUUAAU 128  997-1017 ATUAAGTUGACTAGACACUUUUU 521 995-1017 AD-1725058 AAGUGUCUAGUCAACUUAAUU 129  998-1018AAUUAAGUUGACUAGACACUUUU 522  996-1018 AD-1725059 AGUGUCUAGUCAACUUAAUUU130  999-1019 AAAUTAAGUUGACUAGACACUUU 523  997-1019 AD-1725060GUGUCUAGUCAACUUAAUUGU 131 1000-1020 ACAATUAAGUUGACUAGACACUU 524 998-1020 AD-1725061 UGUCUAGUCAACUUAAUUGAU 132 1001-1021ATCAAUTAAGUTGACUAGACACU 525  999-1021 AD-1725062 GUCUAGUCAACUUAAUUGAGU133 1002-1022 ACUCAATUAAGTUGACUAGACAC 526 1000-1022 AD-1725066AGUCAACUUAAUUGAGAAGGU 134 1006-1026 ACCUTCTCAAUTAAGUUGACUAG 5271004-1026 AD-1725074 UAAUUGAGAAGGUGGCAAGUU 135 1014-1034AACUTGCCACCTUCUCAAUUAAG 528 1012-1034 AD-1725075 AAUUGAGAAGGUGGCAAGUUU136 1015-1035 AAACTUGCCACCUUCUCAAUUAA 529 1013-1035 AD-1725079GAGAAGGUGGCAAGUUAUGGU 137 1019-1039 ACCATAACUUGCCACCUUCUCAA 5301017-1039 AD-1725080 AGAAGGUGGCAAGUUAUGGUU 138 1020-1040AACCAUAACUUGCCACCUUCUCA 531 1018-1040 AD-1725082 AAGGUGGCAAGUUAUGGUGUU139 1022-1042 AACACCAUAACTUGCCACCUUCU 532 1020-1042 AD-1725083AGGUGGCAAGUUAUGGUGUGU 140 1023-1043 ACACACCAUAACUUGCCACCUUC 5331021-1043 AD-1725088 GCAAGUUAUGGUGUGAAGCCU 141 1028-1048AGGCTUCACACCAUAACUUGCCA 534 1026-1048 AD-1725092 GUUAUGGUGUGAAGCCAAGAU142 1032-1052 ATCUTGGCUUCACACCAUAACUU 535 1030-1052 AD-1725095AUGGUGUGAAGCCAAGAUAUU 143 1035-1055 AAUATCTUGGCTUCACACCAUAA 5361033-1055 AD-1725096 UGGUGUGAAGCCAAGAUAUGU 144 1036-1056ACAUAUCUUGGCUUCACACCAUA 537 1034-1056 AD-1725122 AAAAUUUGGGUCAAAGUGUCU145 1082-1102 AGACACTUUGACCCAAAUUUUGG 538 1080-1102 AD-1725123AAAUUUGGGUCAAAGUGUCUU 146 1083-1103 AAGACACUUUGACCCAAAUUUUG 5391081-1103 AD-1725125 AUUUGGGUCAAAGUGUCUGAU 147 1085-1105ATCAGACACUUTGACCCAAAUUU 540 1083-1105 AD-1725156 GUAAUGCAGACUGGGUCACGU148 1116-1136 ACGUGACCCAGTCUGCAUUACUG 541 1114-1136 AD-1725157UAAUGCAGACUGGGUCACGAU 149 1117-1137 ATCGTGACCCAGUCUGCAUUACU 5421115-1137 AD-1725158 AAUGCAGACUGGGUCACGAAU 150 1118-1138ATUCGUGACCCAGUCUGCAUUAC 543 1116-1138 AD-1725159 AUGCAGACUGGGUCACGAAGU151 1119-1139 ACUUCGTGACCCAGUCUGCAUUA 544 1117-1139 AD-1725184AAUGAAAUCAAUUAUGAAGAU 152 1145-1165 ATCUTCAUAAUTGAUUUCAUUGA 5451143-1165 AD-1725186 UGAAAUCAAUUAUGAAGACCU 153 1147-1167AGGUCUTCAUAAUUGAUUUCAUU 546 1145-1167 AD-1725189 AAUCAAUUAUGAAGACCACAU154 1150-1170 ATGUGGTCUUCAUAAUUGAUUUC 547 1148-1170 AD-1725190AUCAAUUAUGAAGACCACAAU 155 1151-1171 ATUGTGGUCUUCAUAAUUGAUUU 5481149-1171 AD-1725191 UCAAUUAUGAAGACCACAAGU 156 1152-1172ACUUGUGGUCUTCAUAAUUGAUU 549 1150-1172 AD-1725192 CAAUUAUGAAGACCACAAGUU157 1153-1173 AACUTGTGGUCTUCAUAAUUGAU 550 1151-1173 AD-1725193AAUUAUGAAGACCACAAGUUU 158 1154-1174 AAACTUGUGGUCUUCAUAAUUGA 5511152-1174 AD-1725194 AUUAUGAAGACCACAAGUUGU 159 1155-1175ACAACUTGUGGTCUUCAUAAUUG 552 1153-1175 AD-1725195 UUAUGAAGACCACAAGUUGAU160 1156-1176 ATCAACTUGUGGUCUUCAUAAUU 553 1154-1176 AD-1725196UAUGAAGACCACAAGUUGAAU 161 1157-1177 ATUCAACUUGUGGUCUUCAUAAU 5541155-1177 AD-1725197 AUGAAGACCACAAGUUGAAGU 162 1158-1178ACUUCAACUUGTGGUCUUCAUAA 555 1156-1178 AD-1725198 UGAAGACCACAAGUUGAAGUU163 1159-1179 AACUTCAACUUGUGGUCUUCAUA 556 1157-1179 AD-1725199GAAGACCACAAGUUGAAGUCU 164 1160-1180 AGACTUCAACUTGUGGUCUUCAU 5571158-1180 AD-1725200 AAGACCACAAGUUGAAGUCAU 165 1161-1181ATGACUTCAACTUGUGGUCUUCA 558 1159-1181 AD-1725201 AGACCACAAGUUGAAGUCAGU166 1162-1182 ACUGACTUCAACUUGUGGUCUUC 559 1160-1182 AD-1725203ACCACAAGUUGAAGUCAGGGU 167 1164-1184 ACCCTGACUUCAACUUGUGGUCU 5601162-1184 AD-1725204 CCACAAGUUGAAGUCAGGGAU 168 1165-1185ATCCCUGACUUCAACUUGUGGUC 561 1163-1185 AD-1725205 CACAAGUUGAAGUCAGGGACU169 1166-1186 AGUCCCTGACUTCAACUUGUGGU 562 1164-1186 AD-1725206ACAAGUUGAAGUCAGGGACUU 170 1167-1187 AAGUCCCUGACTUCAACUUGUGG 5631165-1187 AD-1725208 AAGUUGAAGUCAGGGACUAAU 171 1169-1189ATUAGUCCCUGACUUCAACUUGU 564 1167-1189 AD-1725211 UUGAAGUCAGGGACUAACACU172 1172-1192 AGUGTUAGUCCCUGACUUCAACU 565 1170-1192 AD-1725212UGAAGUCAGGGACUAACACCU 173 1173-1193 AGGUGUTAGUCCCUGACUUCAAC 5661171-1193 AD-1725215 AGUCAGGGACUAACACCAAGU 174 1176-1196ACUUGGTGUUAGUCCCUGACUUC 567 1174-1196 AD-1725216 GUCAGGGACUAACACCAAGAU175 1177-1197 ATCUTGGUGUUAGUCCCUGACUU 568 1175-1197 AD-1725243CCAGGCAGUGUACAGCAUGAU 176 1204-1224 ATCATGCUGUACACUGCCUGGAG 5691202-1224 AD-1725244 CAGGCAGUGUACAGCAUGAUU 177 1205-1225AAUCAUGCUGUACACUGCCUGGA 570 1203-1225 AD-1725245 AGGCAGUGUACAGCAUGAUGU178 1206-1226 ACAUCATGCUGTACACUGCCUGG 571 1204-1226 AD-1725247GCAGUGUACAGCAUGAUGAGU 179 1208-1228 ACUCAUCAUGCTGUACACUGCCU 5721206-1228 AD-1725327 CUGAUGGAUUGCACAACAUGU 180 1290-1310ACAUGUTGUGCAAUCCAUCAGUC 573 1288-1310 AD-1725328 UGAUGGAUUGCACAACAUGGU181 1291-1311 ACCATGTUGUGCAAUCCAUCAGU 574 1289-1311 AD-1725329GAUGGAUUGCACAACAUGGGU 182 1292-1312 ACCCAUGUUGUGCAAUCCAUCAG 5751290-1312 AD-1725330 AUGGAUUGCACAACAUGGGCU 183 1293-1313AGCCCATGUUGTGCAAUCCAUCA 576 1291-1313 AD-1725331 UGGAUUGCACAACAUGGGCGU184 1294-1314 ACGCCCAUGUUGUGCAAUCCAUC 577 1292-1314 AD-1725332GGAUUGCACAACAUGGGCGGU 185 1295-1315 ACCGCCCAUGUTGUGCAAUCCAU 5781293-1315 AD-1725333 GACCCAAUUACUGUCAUUGAU 186 1316-1336ATCAAUGACAGTAAUUGGGUCCC 579 1314-1336 AD-1725334 ACCCAAUUACUGUCAUUGAUU187 1317-1337 AAUCAATGACAGUAAUUGGGUCC 580 1315-1337 AD-1725336CCAAUUACUGUCAUUGAUGAU 188 1319-1339 ATCATCAAUGACAGUAAUUGGGU 5811317-1339 AD-1725344 UGUCAUUGAUGAGAUCCGGGU 189 1327-1347ACCCGGAUCUCAUCAAUGACAGU 582 1325-1347 AD-1725345 GUCAUUGAUGAGAUCCGGGAU190 1328-1348 ATCCCGGAUCUCAUCAAUGACAG 583 1326-1348 AD-1725347CAUUGAUGAGAUCCGGGACUU 191 1330-1350 AAGUCCCGGAUCUCAUCAAUGAC 5841328-1350 AD-1725348 AUUGAUGAGAUCCGGGACUUU 192 1331-1351AAAGTCCCGGATCUCAUCAAUGA 585 1329-1351 AD-1725376 UUGGCAAGGAUCGCAAAAACU193 1359-1379 AGUUTUTGCGATCCUUGCCAAUG 586 1357-1379 AD-1725377UGGCAAGGAUCGCAAAAACCU 194 1360-1380 AGGUTUTUGCGAUCCUUGCCAAU 5871358-1380 AD-1725378 GGCAAGGAUCGCAAAAACCCU 195 1361-1381AGGGTUTUUGCGAUCCUUGCCAA 588 1359-1381 AD-1725397 CAAGGGAGGAUUAUCUGGAUU196 1380-1400 AAUCCAGAUAATCCUCCCUUGGG 589 1378-1400 AD-1725402GAGGAUUAUCUGGAUGUCUAU 197 1385-1405 ATAGACAUCCAGAUAAUCCUCCC 5901383-1405 AD-1725403 AGGAUUAUCUGGAUGUCUAUU 198 1386-1406AAUAGACAUCCAGAUAAUCCUCC 591 1384-1406 AD-1725404 GGAUUAUCUGGAUGUCUAUGU199 1387-1407 ACAUAGACAUCCAGAUAAUCCUC 592 1385-1407 AD-1725405GAUUAUCUGGAUGUCUAUGUU 200 1388-1408 AACATAGACAUCCAGAUAAUCCU 5931386-1408 AD-1725406 AUUAUCUGGAUGUCUAUGUGU 201 1389-1409ACACAUAGACATCCAGAUAAUCC 594 1387-1409 AD-1725407 UUAUCUGGAUGUCUAUGUGUU202 1390-1410 AACACATAGACAUCCAGAUAAUC 595 1388-1410 AD-1725408UAUCUGGAUGUCUAUGUGUUU 203 1391-1411 AAACACAUAGACAUCCAGAUAAU 5961389-1411 AD-1725409 AUCUGGAUGUCUAUGUGUUUU 204 1392-1412AAAACACAUAGACAUCCAGAUAA 597 1390-1412 AD-1725410 UCUGGAUGUCUAUGUGUUUGU205 1393-1413 ACAAACACAUAGACAUCCAGAUA 598 1391-1413 AD-1725411CUGGAUGUCUAUGUGUUUGGU 206 1394-1414 ACCAAACACAUAGACAUCCAGAU 5991392-1414 AD-1725427 AACCAAGUGAACAUCAAUGCU 207 1430-1450AGCATUGAUGUTCACUUGGUUCA 600 1428-1450 AD-1725428 ACCAAGUGAACAUCAAUGCUU208 1431-1451 AAGCAUTGAUGTUCACUUGGUUC 601 1429-1451 AD-1725429CCAAGUGAACAUCAAUGCUUU 209 1432-1452 AAAGCATUGAUGUUCACUUGGUU 6021430-1452 AD-1725430 CAAGUGAACAUCAAUGCUUUU 210 1433-1453AAAAGCAUUGATGUUCACUUGGU 603 1431-1453 AD-1725439 AUCAAUGCUUUGGCUUCCAAU211 1442-1462 ATUGGAAGCCAAAGCAUUGAUGU 604 1440-1462 AD-1725440UCAAUGCUUUGGCUUCCAAGU 212 1443-1463 ACUUGGAAGCCAAAGCAUUGAUG 6051441-1463 AD-1725441 CAAUGCUUUGGCUUCCAAGAU 213 1444-1464ATCUTGGAAGCCAAAGCAUUGAU 606 1442-1464 AD-1725449 UGGCUUCCAAGAAAGACAAUU214 1452-1472 AAUUGUCUUUCTUGGAAGCCAAA 607 1450-1472 AD-1725453UUCCAAGAAAGACAAUGAGCU 215 1456-1476 AGCUCATUGUCTUUCUUGGAAGC 6081454-1476 AD-1725454 UCCAAGAAAGACAAUGAGCAU 216 1457-1477ATGCTCAUUGUCUUUCUUGGAAG 609 1455-1477 AD-1725456 CAAGAAAGACAAUGAGCAACU217 1459-1479 AGUUGCTCAUUGUCUUUCUUGGA 610 1457-1479 AD-1725457AAGAAAGACAAUGAGCAACAU 218 1460-1480 ATGUTGCUCAUTGUCUUUCUUGG 6111458-1480 AD-1725460 AAAGACAAUGAGCAACAUGUU 219 1463-1483AACATGTUGCUCAUUGUCUUUCU 612 1461-1483 AD-1725462 AGACAAUGAGCAACAUGUGUU220 1465-1485 AACACATGUUGCUCAUUGUCUUU 613 1463-1485 AD-1725463GACAAUGAGCAACAUGUGUUU 221 1466-1486 AAACACAUGUUGCUCAUUGUCUU 6141464-1486 AD-1725464 ACAAUGAGCAACAUGUGUUCU 222 1467-1487AGAACACAUGUTGCUCAUUGUCU 615 1465-1487 AD-1725465 CAAUGAGCAACAUGUGUUCAU223 1468-1488 ATGAACACAUGTUGCUCAUUGUC 616 1466-1488 AD-1725467AUGAGCAACAUGUGUUCAAAU 224 1470-1490 ATUUGAACACATGUUGCUCAUUG 6171468-1490 AD-1725469 GAGCAACAUGUGUUCAAAGUU 225 1472-1492AACUTUGAACACAUGUUGCUCAU 618 1470-1492 AD-1725470 AGCAACAUGUGUUCAAAGUCU226 1473-1493 AGACTUTGAACACAUGUUGCUCA 619 1471-1493 AD-1725472CAACAUGUGUUCAAAGUCAAU 227 1475-1495 ATUGACTUUGAACACAUGUUGCU 6201473-1495 AD-1725473 AACAUGUGUUCAAAGUCAAGU 228 1476-1496ACUUGACUUUGAACACAUGUUGC 621 1474-1496 AD-1725474 ACAUGUGUUCAAAGUCAAGGU229 1477-1497 ACCUTGACUUUGAACACAUGUUG 622 1475-1497 AD-1725476AUGUGUUCAAAGUCAAGGAUU 230 1479-1499 AAUCCUTGACUTUGAACACAUGU 6231477-1499 AD-1725477 UGUGUUCAAAGUCAAGGAUAU 231 1480-1500ATAUCCTUGACTUUGAACACAUG 624 1478-1500 AD-1725478 GUGUUCAAAGUCAAGGAUAUU232 1481-1501 AAUATCCUUGACUUUGAACACAU 625 1479-1501 AD-1725481UUCAAAGUCAAGGAUAUGGAU 233 1484-1504 ATCCAUAUCCUTGACUUUGAACA 6261482-1504 AD-1725482 UCAAAGUCAAGGAUAUGGAAU 234 1485-1505ATUCCATAUCCTUGACUUUGAAC 627 1483-1505 AD-1725483 CAAAGUCAAGGAUAUGGAAAU235 1486-1506 ATUUCCAUAUCCUUGACUUUGAA 628 1484-1506 AD-1725534UGAAAGCCAGUCUCUGAGUCU 236 1537-1557 AGACTCAGAGACUGGCUUUCAUC 6291535-1557 AD-1725535 GAAAGCCAGUCUCUGAGUCUU 237 1538-1558AAGACUCAGAGACUGGCUUUCAU 630 1536-1558 AD-1725548 UGAGUCUCUGUGGCAUGGUUU238 1551-1571 AAACCATGCCACAGAGACUCAGA 631 1549-1571 AD-1725552UCUCUGUGGCAUGGUUUGGGU 239 1555-1575 ACCCAAACCAUGCCACAGAGACU 6321553-1575 AD-1725556 UGUGGCAUGGUUUGGGAACAU 240 1559-1579ATGUTCCCAAACCAUGCCACAGA 633 1557-1579 AD-1725558 UGGCAUGGUUUGGGAACACAU241 1561-1581 ATGUGUTCCCAAACCAUGCCACA 634 1559-1581 AD-1725580AAGGGUACCGAUUACCACAAU 242 1583-1603 ATUGTGGUAAUCGGUACCCUUCC 6351581-1603 AD-1725582 GGGUACCGAUUACCACAAGCU 243 1585-1605AGCUTGTGGUAAUCGGUACCCUU 636 1583-1605 AD-1725585 UACCGAUUACCACAAGCAACU244 1588-1608 AGUUGCTUGUGGUAAUCGGUACC 637 1586-1608 AD-1725587CCGAUUACCACAAGCAACCAU 245 1590-1610 ATGGTUGCUUGTGGUAAUCGGUA 6381588-1610 AD-1725588 CGAUUACCACAAGCAACCAUU 246 1591-1611AAUGGUTGCUUGUGGUAAUCGGU 639 1589-1611 AD-1725590 AUUACCACAAGCAACCAUGGU247 1593-1613 ACCATGGUUGCTUGUGGUAAUCG 640 1591-1613 AD-1725591UUACCACAAGCAACCAUGGCU 248 1594-1614 AGCCAUGGUUGCUUGUGGUAAUC 6411592-1614 AD-1725592 UACCACAAGCAACCAUGGCAU 249 1595-1615ATGCCATGGUUGCUUGUGGUAAU 642 1593-1615 AD-1725593 ACCACAAGCAACCAUGGCAGU250 1596-1616 ACUGCCAUGGUTGCUUGUGGUAA 643 1594-1616 AD-1725598AAGCAACCAUGGCAGGCCAAU 251 1601-1621 ATUGGCCUGCCAUGGUUGCUUGU 6441599-1621 AD-1725603 ACCAUGGCAGGCCAAGAUCUU 252 1606-1626AAGATCTUGGCCUGCCAUGGUUG 645 1604-1626 AD-1725604 CCAUGGCAGGCCAAGAUCUCU253 1607-1627 AGAGAUCUUGGCCUGCCAUGGUU 646 1605-1627 AD-1725605CAUGGCAGGCCAAGAUCUCAU 254 1608-1628 ATGAGATCUUGGCCUGCCAUGGU 6471606-1628 AD-1725643 GCUGUGGUGUCUGAGUACUUU 255 1667-1687AAAGTACUCAGACACCACAGCCC 648 1665-1687 AD-1725644 CUGUGGUGUCUGAGUACUUUU256 1668-1688 AAAAGUACUCAGACACCACAGCC 649 1666-1688 AD-1725645UGUGGUGUCUGAGUACUUUGU 257 1669-1689 ACAAAGTACUCAGACACCACAGC 6501667-1689 AD-1725646 GUGGUGUCUGAGUACUUUGUU 258 1670-1690AACAAAGUACUCAGACACCACAG 651 1668-1690 AD-1725647 UGGUGUCUGAGUACUUUGUGU259 1671-1691 ACACAAAGUACTCAGACACCACA 652 1669-1691 AD-1725667CUGACAGCAGCACAUUGUUUU 260 1691-1711 AAAACAAUGUGCUGCUGUCAGCA 6531689-1711 AD-1725716 AAGCGGGACCUGGAGAUAGAU 261 1760-1780ATCUAUCUCCAGGUCCCGCUUCU 654 1758-1780 AD-1725717 AGCGGGACCUGGAGAUAGAAU262 1761-1781 ATUCTATCUCCAGGUCCCGCUUC 655 1759-1781 AD-1725756GAAGCAGGAAUUCCUGAAUUU 263 1823-1843 AAAUTCAGGAATUCCUGCUUCUU 6561821-1843 AD-1725757 AAGCAGGAAUUCCUGAAUUUU 264 1824-1844AAAATUCAGGAAUUCCUGCUUCU 657 1822-1844 AD-1725759 GCAGGAAUUCCUGAAUUUUAU265 1826-1846 ATAAAATUCAGGAAUUCCUGCUU 658 1824-1846 AD-1725760CAGGAAUUCCUGAAUUUUAUU 266 1827-1847 AAUAAAAUUCAGGAAUUCCUGCU 6591825-1847 AD-1725761 AGGAAUUCCUGAAUUUUAUGU 267 1828-1848ACAUAAAAUUCAGGAAUUCCUGC 660 1826-1848 AD-1725762 GGAAUUCCUGAAUUUUAUGAU268 1829-1849 ATCATAAAAUUCAGGAAUUCCUG 661 1827-1849 AD-1725763GAAUUCCUGAAUUUUAUGACU 269 1830-1850 AGUCAUAAAAUTCAGGAAUUCCU 6621828-1850 AD-1725764 AAUUCCUGAAUUUUAUGACUU 270 1831-1851AAGUCATAAAATUCAGGAAUUCC 663 1829-1851 AD-1725765 AUUCCUGAAUUUUAUGACUAU271 1832-1852 ATAGTCAUAAAAUUCAGGAAUUC 664 1830-1852 AD-1725766UUCCUGAAUUUUAUGACUAUU 272 1833-1853 AAUAGUCAUAAAAUUCAGGAAUU 6651831-1853 AD-1725767 UCCUGAAUUUUAUGACUAUGU 273 1834-1854ACAUAGTCAUAAAAUUCAGGAAU 666 1832-1854 AD-1725768 CCUGAAUUUUAUGACUAUGAU274 1835-1855 ATCATAGUCAUAAAAUUCAGGAA 667 1833-1855 AD-1725769CUGAAUUUUAUGACUAUGACU 275 1836-1856 AGUCAUAGUCATAAAAUUCAGGA 6681834-1856 AD-1725771 GAAUUUUAUGACUAUGACGUU 276 1838-1858AACGTCAUAGUCAUAAAAUUCAG 669 1836-1858 AD-1725772 AAUUUUAUGACUAUGACGUUU277 1839-1859 AAACGUCAUAGTCAUAAAAUUCA 670 1837-1859 AD-1725773AUUUUAUGACUAUGACGUUGU 278 1840-1860 ACAACGTCAUAGUCAUAAAAUUC 6711838-1860 AD-1725775 UUUAUGACUAUGACGUUGCCU 279 1842-1862AGGCAACGUCATAGUCAUAAAAU 672 1840-1862 AD-1725776 UUAUGACUAUGACGUUGCCCU280 1843-1863 AGGGCAACGUCAUAGUCAUAAAA 673 1841-1863 AD-1725777UAUGACUAUGACGUUGCCCUU 281 1844-1864 AAGGGCAACGUCAUAGUCAUAAA 6741842-1864 AD-1725778 AUGACUAUGACGUUGCCCUGU 282 1845-1865ACAGGGCAACGTCAUAGUCAUAA 675 1843-1865 AD-1725779 UGACUAUGACGUUGCCCUGAU283 1846-1866 ATCAGGGCAACGUCAUAGUCAUA 676 1844-1866 AD-1725780GACUAUGACGUUGCCCUGAUU 284 1847-1867 AAUCAGGGCAACGUCAUAGUCAU 6771845-1867 AD-1725784 AUGACGUUGCCCUGAUCAAGU 285 1851-1871ACUUGATCAGGGCAACGUCAUAG 678 1849-1871 AD-1725785 UGACGUUGCCCUGAUCAAGCU286 1852-1872 AGCUTGAUCAGGGCAACGUCAUA 679 1850-1872 AD-1725786GACGUUGCCCUGAUCAAGCUU 287 1853-1873 AAGCTUGAUCAGGGCAACGUCAU 6801851-1873 AD-1725787 ACGUUGCCCUGAUCAAGCUCU 288 1854-1874AGAGCUTGAUCAGGGCAACGUCA 681 1852-1874 AD-1725789 GUUGCCCUGAUCAAGCUCAAU289 1856-1876 ATUGAGCUUGATCAGGGCAACGU 682 1854-1876 AD-1725790UUGCCCUGAUCAAGCUCAAGU 290 1857-1877 ACUUGAGCUUGAUCAGGGCAACG 6831855-1877 AD-1725828 CAGACUAUCAGGCCCAUUUGU 291 1895-1915ACAAAUGGGCCTGAUAGUCUGGC 684 1893-1915 AD-1725829 AGACUAUCAGGCCCAUUUGUU292 1896-1916 AACAAATGGGCCUGAUAGUCUGG 685 1894-1916 AD-1725830GACUAUCAGGCCCAUUUGUCU 293 1897-1917 AGACAAAUGGGCCUGAUAGUCUG 6861895-1917 AD-1725831 ACUAUCAGGCCCAUUUGUCUU 294 1898-1918AAGACAAAUGGGCCUGAUAGUCU 687 1896-1918 AD-1725832 CUAUCAGGCCCAUUUGUCUCU295 1899-1919 AGAGACAAAUGGGCCUGAUAGUC 688 1897-1919 AD-1725840CGAGGGAACAACUCGAGCUUU 296 1927-1947 AAAGCUCGAGUTGUUCCCUCGGU 6891925-1947 AD-1725841 GAGGGAACAACUCGAGCUUUU 297 1928-1948AAAAGCTCGAGTUGUUCCCUCGG 690 1926-1948 AD-1725842 AGGGAACAACUCGAGCUUUGU298 1929-1949 ACAAAGCUCGAGUUGUUCCCUCG 691 1927-1949 AD-1725845GAACAACUCGAGCUUUGAGGU 299 1932-1952 ACCUCAAAGCUCGAGUUGUUCCC 6921930-1952 AD-1725846 AACAACUCGAGCUUUGAGGCU 300 1933-1953AGCCTCAAAGCTCGAGUUGUUCC 693 1931-1953 AD-1725848 CAACUCGAGCUUUGAGGCUUU301 1935-1955 AAAGCCTCAAAGCUCGAGUUGUU 694 1933-1955 AD-1725849AACUCGAGCUUUGAGGCUUCU 302 1936-1956 AGAAGCCUCAAAGCUCGAGUUGU 6951934-1956 AD-1725850 ACUCGAGCUUUGAGGCUUCCU 303 1937-1957AGGAAGCCUCAAAGCUCGAGUUG 696 1935-1957 AD-1725854 GAGCUUUGAGGCUUCCUCCAU304 1941-1961 ATGGAGGAAGCCUCAAAGCUCGA 697 1939-1961 AD-1725855AGCUUUGAGGCUUCCUCCAAU 305 1942-1962 ATUGGAGGAAGCCUCAAAGCUCG 6981940-1962 AD-1725856 GCUUUGAGGCUUCCUCCAACU 306 1943-1963AGUUGGAGGAAGCCUCAAAGCUC 699 1941-1963 AD-1725857 CUUUGAGGCUUCCUCCAACUU307 1944-1964 AAGUTGGAGGAAGCCUCAAAGCU 700 1942-1964 AD-1725858UUUGAGGCUUCCUCCAACUAU 308 1945-1965 ATAGTUGGAGGAAGCCUCAAAGC 7011943-1965 AD-1725861 GAGGCUUCCUCCAACUACCAU 309 1948-1968ATGGTAGUUGGAGGAAGCCUCAA 702 1946-1968 AD-1725862 AGGCUUCCUCCAACUACCACU310 1949-1969 AGUGGUAGUUGGAGGAAGCCUCA 703 1947-1969 AD-1725864GCUUCCUCCAACUACCACUUU 311 1951-1971 AAAGTGGUAGUTGGAGGAAGCCU 7041949-1971 AD-1725866 UUCCUCCAACUACCACUUGCU 312 1953-1973AGCAAGTGGUAGUUGGAGGAAGC 705 1951-1973 AD-1725867 UCCUCCAACUACCACUUGCCU313 1954-1974 AGGCAAGUGGUAGUUGGAGGAAG 706 1952-1974 AD-1725872CAACUACCACUUGCCAGCAAU 314 1959-1979 ATUGCUGGCAAGUGGUAGUUGGA 7071957-1979 AD-1725874 ACUACCACUUGCCAGCAACAU 315 1961-1981ATGUTGCUGGCAAGUGGUAGUUG 708 1959-1981 AD-1725907 CUCCCUGCACAGGAUAUCAAU316 1994-2014 ATUGAUAUCCUGUGCAGGGAGCA 709 1992-2014 AD-1725908UCCCUGCACAGGAUAUCAAAU 317 1995-2015 ATUUGATAUCCTGUGCAGGGAGC 7101993-2015 AD-1725909 CCCUGCACAGGAUAUCAAAGU 318 1996-2016ACUUTGAUAUCCUGUGCAGGGAG 711 1994-2016 AD-1725911 CUGCACAGGAUAUCAAAGCUU319 1998-2018 AAGCTUTGAUATCCUGUGCAGGG 712 1996-2018 AD-1725916CAGGAUAUCAAAGCUCUGUUU 320 2003-2023 AAACAGAGCUUTGAUAUCCUGUG 7132001-2023 AD-1725919 GAUAUCAAAGCUCUGUUUGUU 321 2006-2026AACAAACAGAGCUUUGAUAUCCU 714 2004-2026 AD-1725925 AAAGCUCUGUUUGUGUCUGAU322 2012-2032 ATCAGACACAAACAGAGCUUUGA 715 2010-2032 AD-1725957GCUGACUCGGAAGGAGGUCUU 323 2044-2064 AAGACCTCCUUCCGAGUCAGCUU 7162042-2064 AD-1725958 CUGACUCGGAAGGAGGUCUAU 324 2045-2065ATAGACCUCCUTCCGAGUCAGCU 717 2043-2065 AD-1725961 ACUCGGAAGGAGGUCUACAUU325 2048-2068 AAUGTAGACCUCCUUCCGAGUCA 718 2046-2068 AD-1725963UCGGAAGGAGGUCUACAUCAU 326 2050-2070 ATGATGTAGACCUCCUUCCGAGU 7192048-2070 AD-1725964 CGGAAGGAGGUCUACAUCAAU 327 2051-2071ATUGAUGUAGACCUCCUUCCGAG 720 2049-2071 AD-1725967 AAGGAGGUCUACAUCAAGAAU328 2054-2074 ATUCTUGAUGUAGACCUCCUUCC 721 2052-2074 AD-1725968AGGAGGUCUACAUCAAGAAUU 329 2055-2075 AAUUCUTGAUGTAGACCUCCUUC 7222053-2075 AD-1725974 AAGAAAGGCAGCUGUGAGAGU 330 2081-2101ACUCTCACAGCTGCCUUUCUUAU 723 2079-2101 AD-1725977 AAAGGCAGCUGUGAGAGAGAU331 2084-2104 ATCUCUCUCACAGCUGCCUUUCU 724 2082-2104 AD-1725983AGCUGUGAGAGAGAUGCUCAU 332 2090-2110 ATGAGCAUCUCTCUCACAGCUGC 7252088-2110 AD-1725985 CUGUGAGAGAGAUGCUCAAUU 333 2092-2112AAUUGAGCAUCTCUCUCACAGCU 726 2090-2112 AD-1725986 UGUGAGAGAGAUGCUCAAUAU334 2093-2113 ATAUTGAGCAUCUCUCUCACAGC 727 2091-2113 AD-1725987GUGAGAGAGAUGCUCAAUAUU 335 2094-2114 AAUATUGAGCATCUCUCUCACAG 7282092-2114 AD-1725988 UGAGAGAGAUGCUCAAUAUGU 336 2095-2115ACAUAUTGAGCAUCUCUCUCACA 729 2093-2115 AD-1725989 GAGAGAGAUGCUCAAUAUGCU337 2096-2116 AGCATATUGAGCAUCUCUCUCAC 730 2094-2116 AD-1725991CAGGCUAUGACAAAGUCAAGU 338 2118-2138 ACUUGACUUUGTCAUAGCCUGGG 7312116-2138 AD-1725992 AGGCUAUGACAAAGUCAAGGU 339 2119-2139ACCUTGACUUUGUCAUAGCCUGG 732 2117-2139 AD-1725993 GGCUAUGACAAAGUCAAGGAU340 2120-2140 ATCCTUGACUUTGUCAUAGCCUG 733 2118-2140 AD-1725999GACAAAGUCAAGGACAUCUCU 341 2126-2146 AGAGAUGUCCUTGACUUUGUCAU 7342124-2146 AD-1726014 UCGGUUCCUUUGUACUGGAGU 342 2161-2181ACUCCAGUACAAAGGAACCGAGG 735 2159-2181 AD-1726015 CGGUUCCUUUGUACUGGAGGU343 2162-2182 ACCUCCAGUACAAAGGAACCGAG 736 2160-2182 AD-1726016GGUUCCUUUGUACUGGAGGAU 344 2163-2183 ATCCTCCAGUACAAAGGAACCGA 7372161-2183 AD-1726018 UUCCUUUGUACUGGAGGAGUU 345 2165-2185AACUCCTCCAGTACAAAGGAACC 738 2163-2185 AD-1726020 CCUUUGUACUGGAGGAGUGAU346 2167-2187 ATCACUCCUCCAGUACAAAGGAA 739 2165-2187 AD-1726023UUGUACUGGAGGAGUGAGUCU 347 2170-2190 AGACTCACUCCTCCAGUACAAAG 7402168-2190 AD-1726024 UGUACUGGAGGAGUGAGUCCU 348 2171-2191AGGACUCACUCCUCCAGUACAAA 741 2169-2191 AD-1726025 GUACUGGAGGAGUGAGUCCCU349 2172-2192 AGGGACTCACUCCUCCAGUACAA 742 2170-2192 AD-1726027ACUGGAGGAGUGAGUCCCUAU 350 2174-2194 ATAGGGACUCACUCCUCCAGUAC 7432172-2194 AD-1726029 UGGAGGAGUGAGUCCCUAUGU 351 2176-2196ACAUAGGGACUCACUCCUCCAGU 744 2174-2196 AD-1726031 GAGGAGUGAGUCCCUAUGCUU352 2178-2198 AAGCAUAGGGACUCACUCCUCCA 745 2176-2198 AD-1726033GGAGUGAGUCCCUAUGCUGAU 353 2180-2200 ATCAGCAUAGGGACUCACUCCUC 7462178-2200 AD-1726034 GAGUGAGUCCCUAUGCUGACU 354 2181-2201AGUCAGCAUAGGGACUCACUCCU 747 2179-2201 AD-1726036 CAAUACUUGCAGAGGUGAUUU355 2203-2223 AAAUCACCUCUGCAAGUAUUGGG 748 2201-2223 AD-1726037AAUACUUGCAGAGGUGAUUCU 356 2204-2224 AGAATCACCUCTGCAAGUAUUGG 7492202-2224 AD-1726039 UACUUGCAGAGGUGAUUCUGU 357 2206-2226ACAGAATCACCTCUGCAAGUAUU 750 2204-2226 AD-1726041 CUUGCAGAGGUGAUUCUGGCU358 2208-2228 AGCCAGAAUCACCUCUGCAAGUA 751 2206-2228 AD-1726042UUGCAGAGGUGAUUCUGGCGU 359 2209-2229 ACGCCAGAAUCACCUCUGCAAGU 7522207-2229 AD-1726048 UGAUAGUUCACAAGAGAAGUU 360 2235-2255AACUTCTCUUGTGAACUAUCAAG 753 2233-2255 AD-1726049 GAUAGUUCACAAGAGAAGUCU361 2236-2256 AGACTUCUCUUGUGAACUAUCAA 754 2234-2256 AD-1726050AUAGUUCACAAGAGAAGUCGU 362 2237-2257 ACGACUTCUCUTGUGAACUAUCA 7552235-2257 AD-1726051 UAGUUCACAAGAGAAGUCGUU 363 2238-2258AACGACTUCUCTUGUGAACUAUC 756 2236-2258 AD-1726052 AGUUCACAAGAGAAGUCGUUU364 2239-2259 AAACGACUUCUCUUGUGAACUAU 757 2237-2259 AD-1726053GUUCACAAGAGAAGUCGUUUU 365 2240-2260 AAAACGACUUCTCUUGUGAACUA 7582238-2260 AD-1726054 UUCACAAGAGAAGUCGUUUCU 366 2241-2261AGAAACGACUUCUCUUGUGAACU 759 2239-2261 AD-1726055 UCACAAGAGAAGUCGUUUCAU367 2242-2262 ATGAAACGACUTCUCUUGUGAAC 760 2240-2262 AD-1726056CACAAGAGAAGUCGUUUCAUU 368 2243-2263 AAUGAAACGACTUCUCUUGUGAA 7612241-2263 AD-1726057 ACAAGAGAAGUCGUUUCAUUU 369 2244-2264AAAUGAAACGACUUCUCUUGUGA 762 2242-2264 AD-1726058 CAAGAGAAGUCGUUUCAUUCU370 2245-2265 AGAATGAAACGACUUCUCUUGUG 763 2243-2265 AD-1726059AAGAGAAGUCGUUUCAUUCAU 371 2246-2266 ATGAAUGAAACGACUUCUCUUGU 7642244-2266 AD-1726060 AGAGAAGUCGUUUCAUUCAAU 372 2247-2267ATUGAATGAAACGACUUCUCUUG 765 2245-2267 AD-1726061 GAGAAGUCGUUUCAUUCAAGU373 2248-2268 ACUUGAAUGAAACGACUUCUCUU 766 2246-2268 AD-1726062AGAAGUCGUUUCAUUCAAGUU 374 2249-2269 AACUTGAAUGAAACGACUUCUCU 7672247-2269 AD-1726063 GAAGUCGUUUCAUUCAAGUUU 375 2250-2270AAACTUGAAUGAAACGACUUCUC 768 2248-2270 AD-1726064 AAGUCGUUUCAUUCAAGUUGU376 2251-2271 ACAACUTGAAUGAAACGACUUCU 769 2249-2271 AD-1726065AGUCGUUUCAUUCAAGUUGGU 377 2252-2272 ACCAACTUGAATGAAACGACUUC 7702250-2272 AD-1726079 GAGUAGUGGAUGUCUGCAAAU 378 2286-2306ATUUGCAGACATCCACUACUCCC 771 2284-2306 AD-1726080 AGUAGUGGAUGUCUGCAAAAU379 2287-2307 ATUUTGCAGACAUCCACUACUCC 772 2285-2307 AD-1726081GUAGUGGAUGUCUGCAAAAAU 380 2288-2308 ATUUTUGCAGACAUCCACUACUC 7732286-2308 AD-1726082 UAGUGGAUGUCUGCAAAAACU 381 2289-2309AGUUTUTGCAGACAUCCACUACU 774 2287-2309 AD-1726083 AGUGGAUGUCUGCAAAAACCU382 2290-2310 AGGUTUTUGCAGACAUCCACUAC 775 2288-2310 AD-1726084GUGGAUGUCUGCAAAAACCAU 383 2291-2311 ATGGTUTUUGCAGACAUCCACUA 7762289-2311 AD-1726085 UGGAUGUCUGCAAAAACCAGU 384 2292-2312ACUGGUTUUUGCAGACAUCCACU 777 2290-2312 AD-1726086 GGAUGUCUGCAAAAACCAGAU385 2293-2313 ATCUGGTUUUUGCAGACAUCCAC 778 2291-2313 AD-1726087GAUGUCUGCAAAAACCAGAAU 386 2294-2314 ATUCTGGUUUUTGCAGACAUCCA 7792292-2314 AD-1726090 GUCUGCAAAAACCAGAAGCGU 387 2297-2317ACGCTUCUGGUTUUUGCAGACAU 780 2295-2317 AD-1726091 UCUGCAAAAACCAGAAGCGGU388 2298-2318 ACCGCUTCUGGTUUUUGCAGACA 781 2296-2318 AD-1726092CUGCAAAAACCAGAAGCGGCU 389 2299-2319 AGCCGCTUCUGGUUUUUGCAGAC 7822297-2319 AD-1726095 CAAAAACCAGAAGCGGCAAAU 390 2302-2322ATUUGCCGCUUCUGGUUUUUGCA 783 2300-2322 AD-1726096 AAAAACCAGAAGCGGCAAAAU391 2303-2323 ATUUTGCCGCUTCUGGUUUUUGC 784 2301-2323 AD-1726097AAAACCAGAAGCGGCAAAAGU 392 2304-2324 ACUUTUGCCGCTUCUGGUUUUUG 7852302-2324 AD-1726098 AAACCAGAAGCGGCAAAAGCU 393 2305-2325AGCUTUTGCCGCUUCUGGUUUUU 786 2303-2325 AD-1726099 AACCAGAAGCGGCAAAAGCAU394 2306-2326 ATGCTUTUGCCGCUUCUGGUUUU 787 2304-2326 AD-1726103AGAAGCGGCAAAAGCAGGUAU 395 2310-2330 ATACCUGCUUUTGCCGCUUCUGG 7882308-2330 AD-1726113 AAAGCAGGUACCUGCUCACGU 396 2320-2340ACGUGAGCAGGTACCUGCUUUUG 789 2318-2340 AD-1726159 CAAGUGCUGCCCUGGCUGAAU397 2366-2386 ATUCAGCCAGGGCAGCACUUGAA 790 2364-2386 AD-1726171UGGCUGAAGGAGAAACUCCAU 398 2378-2398 ATGGAGTUUCUCCUUCAGCCAGG 7912376-2398 AD-1726184 AACUCCAAGAUGAGGAUUUGU 399 2391-2411ACAAAUCCUCATCUUGGAGUUUC 792 2389-2411 AD-1726187 UCCAAGAUGAGGAUUUGGGUU400 2394-2414 AACCCAAAUCCTCAUCUUGGAGU 793 2392-2414 AD-1726189CAAGAUGAGGAUUUGGGUUUU 401 2396-2416 AAAACCCAAAUCCUCAUCUUGGA 7942394-2416 AD-1726191 AGAUGAGGAUUUGGGUUUUCU 402 2398-2418AGAAAACCCAAAUCCUCAUCUUG 795 2396-2418 AD-1726201 GUGGGAUUGAAUUAAAACAGU403 2446-2466 ACUGTUTUAAUTCAAUCCCACGC 796 2444-2466 AD-1726202UGGGAUUGAAUUAAAACAGCU 404 2447-2467 AGCUGUTUUAATUCAAUCCCACG 7972445-2467 AD-1726203 GGGAUUGAAUUAAAACAGCUU 405 2448-2468AAGCTGTUUUAAUUCAAUCCCAC 798 2446-2468 AD-1726206 AUUGAAUUAAAACAGCUGCGU406 2451-2471 ACGCAGCUGUUTUAAUUCAAUCC 799 2449-2471 AD-1726207UUGAAUUAAAACAGCUGCGAU 407 2452-2472 ATCGCAGCUGUTUUAAUUCAAUC 8002450-2472 AD-1726208 UGAAUUAAAACAGCUGCGACU 408 2453-2473AGUCGCAGCUGTUUUAAUUCAAU 801 2451-2473 AD-1726209 GAAUUAAAACAGCUGCGACAU409 2454-2474 ATGUCGCAGCUGUUUUAAUUCAA 802 2452-2474 AD-1726815CUGGCUUCUACCCGUACCCUU 34  469-489 AAGGGUACGGGUAGAAGCCAGAA 803  467-489AD-1726927 CCCUACUACAAUGUGAGUGAU 40  633-653 AUCACUCACAUUGUAGUAGGGAG 804 631-653 AD-1726928 CCUACUACAAUGUGAGUGAUU 41  634-654AAUCACUCACAUUGUAGUAGGGA 805  632-654 AD-1726931 ACUACAAUGUGAGUGAUGAGU 43 637-657 ACUCAUCACUCACAUUGUAGUAG 436  635-657 AD-1726934ACAAUGUGAGUGAUGAGAUCU 44  640-660 AGAUCUCAUCACUCACAUUGUAG 437  638-660AD-1726935 CAAUGUGAGUGAUGAGAUCUU 45  641-661 AAGAUCUCAUCACUCACAUUGUA 806 639-661 AD-1726936 AAUGUGAGUGAUGAGAUCUCU 46  642-662AGAGAUCUCAUCACUCACAUUGU 439  640-662 AD-1726937 AUGUGAGUGAUGAGAUCUCUU 47 643-663 AAGAGAUCUCAUCACUCACAUUG 807  641-663 AD-1726938UGUGAGUGAUGAGAUCUCUUU 48  644-664 AAAGAGAUCUCAUCACUCACAUU 441  642-664AD-1726939 GUGAGUGAUGAGAUCUCUUUU 49  645-665 AAAAGAGAUCUCAUCACUCACAU 442 643-665 AD-1726940 UGAGUGAUGAGAUCUCUUUCU 50  646-666AGAAAGAGAUCUCAUCACUCACA 808  644-666 AD-1726941 GAGUGAUGAGAUCUCUUUCCU 51 647-667 AGGAAAGAGAUCUCAUCACUCAC 444  645-667 AD-1726942AGUGAUGAGAUCUCUUUCCAU 52  648-668 AUGGAAAGAGAUCUCAUCACUCA 809  646-668AD-1726944 UGAUGAGAUCUCUUUCCACUU 54  650-670 AAGUGGAAAGAGAUCUCAUCACU 447 648-670 AD-1726952 UCUCUUUCCACUGCUAUGACU 57  658-678AGUCAUAGCAGUGGAAAGAGAUC 810  656-678 AD-1726961 ACUGCUAUGACGGUUACACUU 60 667-687 AAGUGUAACCGUCAUAGCAGUGG 811  665-687 AD-1727012CAGACAGCGAUCUGUGACAAU 67  738-758 AUUGUCACAGAUCGCUGUCUGCC 812  736-758AD-1727059 CUUGAAGACAGCGUCACCUAU 75  825-845 AUAGGUGACGCUGUCUUCAAGGC 813 823-845 AD-1727140 AAGACUCCUUCAUGUACGACU 84  934-954AGUCGUACAUGAAGGAGUCUUGG 477  932-954 AD-1727142 GACUCCUUCAUGUACGACACU 86 936-956 AGUGUCGUACAUGAAGGAGUCUU 814  934-956 AD-1727181AGAGACCAUAGAAGGAGUCGU 89  995-1015 ACGACUCCUUCUAUGGUCUCUGU 815  993-1015AD-1727183 AGACCAUAGAAGGAGUCGAUU 91  997-1017 AAUCGACUCCUUCUAUGGUCUCU816  995-1017 AD-1727184 GACCAUAGAAGGAGUCGAUGU 92  998-1018ACAUCGACUCCUUCUAUGGUCUC 817  996-1018 AD-1727249 UGAACAUCUACCUGGUGCUAU97 1084-1104 AUAGCACCAGGUAGAUGUUCAUG 818 1082-1104 AD-1727261UGGUGCUAGAUGGAUCAGACU 106 1096-1116 AGUCUGAUCCAUCUAGCACCAGG 8191094-1116 AD-1727263 GUGCUAGAUGGAUCAGACAGU 107 1098-1118ACUGUCUGAUCCAUCUAGCACCA 820 1096-1118 AD-1727275 CAACUUCACAGGAGCCAAAAU110 1130-1150 AUUUUGGCUCCUGUGAAGUUGCU 821 1128-1150 AD-1727276AACUUCACAGGAGCCAAAAAU 111 1131-1151 AUUUUUGGCUCCUGUGAAGUUGC 8221129-1151 AD-1727278 CUUCACAGGAGCCAAAAAGUU 113 1133-1153AACUUUUUGGCUCCUGUGAAGUU 823 1131-1153 AD-1727285 GGAGCCAAAAAGUGUCUAGUU120 1140-1160 AACUAGACACUUUUUGGCUCCUG 824 1138-1160 AD-1727286GAGCCAAAAAGUGUCUAGUCU 121 1141-1161 AGACUAGACACUUUUUGGCUCCU 8251139-1161 AD-1727288 GCCAAAAAGUGUCUAGUCAAU 123 1143-1163AUUGACUAGACACUUUUUGGCUC 826 1141-1163 AD-1727289 CCAAAAAGUGUCUAGUCAACU124 1144-1164 AGUUGACUAGACACUUUUUGGCU 517 1142-1164 AD-1727290CAAAAAGUGUCUAGUCAACUU 125 1145-1165 AAGUUGACUAGACACUUUUUGGC 8271143-1165 AD-1727291 AAAAAGUGUCUAGUCAACUUU 126 1146-1166AAAGUUGACUAGACACUUUUUGG 828 1144-1166 AD-1727292 AAAAGUGUCUAGUCAACUUAU127 1147-1167 AUAAGUUGACUAGACACUUUUUG 829 1145-1167 AD-1727293AAAGUGUCUAGUCAACUUAAU 128 1148-1168 AUUAAGUUGACUAGACACUUUUU 8301146-1168 AD-1727298 GUCUAGUCAACUUAAUUGAGU 133 1153-1173ACUCAAUUAAGUUGACUAGACAC 831 1151-1173 AD-1727310 UAAUUGAGAAGGUGGCAAGUU135 1165-1185 AACUUGCCACCUUCUCAAUUAAG 832 1163-1185 AD-1727318AAGGUGGCAAGUUAUGGUGUU 139 1173-1193 AACACCAUAACUUGCCACCUUCU 8331171-1193 AD-1727324 GCAAGUUAUGGUGUGAAGCCU 141 1179-1199AGGCUUCACACCAUAACUUGCCA 834 1177-1199 AD-1727331 AUGGUGUGAAGCCAAGAUAUU143 1186-1206 AAUAUCUUGGCUUCACACCAUAA 835 1184-1206 AD-1727358AAAAUUUGGGUCAAAGUGUCU 145 1233-1253 AGACACUUUGACCCAAAUUUUGG 8361231-1253 AD-1727359 AAAUUUGGGUCAAAGUGUCUU 146 1234-1254AAGACACUUUGACCCAAAUUUUG 539 1232-1254 AD-1727361 AUUUGGGUCAAAGUGUCUGAU147 1236-1256 AUCAGACACUUUGACCCAAAUUU 837 1234-1256 AD-1727392GUAAUGCAGACUGGGUCACGU 148 1267-1287 ACGUGACCCAGUCUGCAUUACUG 8381265-1287 AD-1727420 AAUGAAAUCAAUUAUGAAGAU 152 1296-1316AUCUUCAUAAUUGAUUUCAUUGA 839 1294-1316 AD-1727427 UCAAUUAUGAAGACCACAAGU156 1303-1323 ACUUGUGGUCUUCAUAAUUGAUU 840 1301-1323 AD-1727428CAAUUAUGAAGACCACAAGUU 157 1304-1324 AACUUGUGGUCUUCAUAAUUGAU 8411302-1324 AD-1727430 AUUAUGAAGACCACAAGUUGU 159 1306-1326ACAACUUGUGGUCUUCAUAAUUG 842 1304-1326 AD-1727431 UUAUGAAGACCACAAGUUGAU160 1307-1327 AUCAACUUGUGGUCUUCAUAAUU 843 1305-1327 AD-1727432UAUGAAGACCACAAGUUGAAU 161 1308-1328 AUUCAACUUGUGGUCUUCAUAAU 8441306-1328 AD-1727433 AUGAAGACCACAAGUUGAAGU 162 1309-1329ACUUCAACUUGUGGUCUUCAUAA 845 1307-1329 AD-1727434 UGAAGACCACAAGUUGAAGUU163 1310-1330 AACUUCAACUUGUGGUCUUCAUA 846 1308-1330 AD-1727435GAAGACCACAAGUUGAAGUCU 164 1311-1331 AGACUUCAACUUGUGGUCUUCAU 8471309-1331 AD-1727436 AAGACCACAAGUUGAAGUCAU 165 1312-1332AUGACUUCAACUUGUGGUCUUCA 848 1310-1332 AD-1727441 CACAAGUUGAAGUCAGGGACU169 1317-1337 AGUCCCUGACUUCAACUUGUGGU 849 1315-1337 AD-1727442ACAAGUUGAAGUCAGGGACUU 170 1318-1338 AAGUCCCUGACUUCAACUUGUGG 8501316-1338 AD-1727481 AGGCAGUGUACAGCAUGAUGU 178 1357-1377ACAUCAUGCUGUACACUGCCUGG 851 1355-1377 AD-1727483 GCAGUGUACAGCAUGAUGAGU179 1359-1379 ACUCAUCAUGCUGUACACUGCCU 852 1357-1379 AD-1727565GAUGGAUUGCACAACAUGGGU 182 1443-1463 ACCCAUGUUGUGCAAUCCAUCAG 5751441-1463 AD-1727566 AUGGAUUGCACAACAUGGGCU 183 1444-1464AGCCCAUGUUGUGCAAUCCAUCA 853 1442-1464 AD-1727568 GGAUUGCACAACAUGGGCGGU185 1446-1466 ACCGCCCAUGUUGUGCAAUCCAU 854 1444-1466 AD-1727569GACCCAAUUACUGUCAUUGAU 186 1467-1487 AUCAAUGACAGUAAUUGGGUCCC 8551465-1487 AD-1727570 ACCCAAUUACUGUCAUUGAUU 187 1468-1488AAUCAAUGACAGUAAUUGGGUCC 856 1466-1488 AD-1727572 CCAAUUACUGUCAUUGAUGAU188 1470-1490 AUCAUCAAUGACAGUAAUUGGGU 857 1468-1490 AD-1727584AUUGAUGAGAUCCGGGACUUU 192 1482-1502 AAAGUCCCGGAUCUCAUCAAUGA 8581480-1502 AD-1727612 UUGGCAAGGAUCGCAAAAACU 193 1510-1530AGUUUUUGCGAUCCUUGCCAAUG 859 1508-1530 AD-1727633 CAAGGGAGGAUUAUCUGGAUU196 1531-1551 AAUCCAGAUAAUCCUCCCUUGGG 860 1529-1551 AD-1727638GAGGAUUAUCUGGAUGUCUAU 197 1536-1556 AUAGACAUCCAGAUAAUCCUCCC 8611534-1556 AD-1727639 AGGAUUAUCUGGAUGUCUAUU 198 1537-1557AAUAGACAUCCAGAUAAUCCUCC 591 1535-1557 AD-1727640 GGAUUAUCUGGAUGUCUAUGU199 1538-1558 ACAUAGACAUCCAGAUAAUCCUC 592 1536-1558 AD-1727641GAUUAUCUGGAUGUCUAUGUU 200 1539-1559 AACAUAGACAUCCAGAUAAUCCU 8621537-1559 AD-1727642 AUUAUCUGGAUGUCUAUGUGU 201 1540-1560ACACAUAGACAUCCAGAUAAUCC 863 1538-1560 AD-1727643 UUAUCUGGAUGUCUAUGUGUU202 1541-1561 AACACAUAGACAUCCAGAUAAUC 864 1539-1561 AD-1727644UAUCUGGAUGUCUAUGUGUUU 203 1542-1562 AAACACAUAGACAUCCAGAUAAU 5961540-1562 AD-1727645 AUCUGGAUGUCUAUGUGUUUU 204 1543-1563AAAACACAUAGACAUCCAGAUAA 597 1541-1563 AD-1727646 UCUGGAUGUCUAUGUGUUUGU205 1544-1564 ACAAACACAUAGACAUCCAGAUA 598 1542-1564 AD-1727663AACCAAGUGAACAUCAAUGCU 207 1581-1601 AGCAUUGAUGUUCACUUGGUUCA 8651579-1601 AD-1727664 ACCAAGUGAACAUCAAUGCUU 208 1582-1602AAGCAUUGAUGUUCACUUGGUUC 866 1580-1602 AD-1727665 CCAAGUGAACAUCAAUGCUUU209 1583-1603 AAAGCAUUGAUGUUCACUUGGUU 867 1581-1603 AD-1727666CAAGUGAACAUCAAUGCUUUU 210 1584-1604 AAAAGCAUUGAUGUUCACUUGGU 8681582-1604 AD-1727675 AUCAAUGCUUUGGCUUCCAAU 211 1593-1613AUUGGAAGCCAAAGCAUUGAUGU 869 1591-1613 AD-1727677 CAAUGCUUUGGCUUCCAAGAU213 1595-1615 AUCUUGGAAGCCAAAGCAUUGAU 870 1593-1615 AD-1727685UGGCUUCCAAGAAAGACAAUU 214 1603-1623 AAUUGUCUUUCUUGGAAGCCAAA 8711601-1623 AD-1727689 UUCCAAGAAAGACAAUGAGCU 215 1607-1627AGCUCAUUGUCUUUCUUGGAAGC 872 1605-1627 AD-1727690 UCCAAGAAAGACAAUGAGCAU216 1608-1628 AUGCUCAUUGUCUUUCUUGGAAG 873 1606-1628 AD-1727693AAGAAAGACAAUGAGCAACAU 218 1611-1631 AUGUUGCUCAUUGUCUUUCUUGG 8741609-1631 AD-1727696 AAAGACAAUGAGCAACAUGUU 219 1614-1634AACAUGUUGCUCAUUGUCUUUCU 875 1612-1634 AD-1727698 AGACAAUGAGCAACAUGUGUU220 1616-1636 AACACAUGUUGCUCAUUGUCUUU 876 1614-1636 AD-1727699GACAAUGAGCAACAUGUGUUU 221 1617-1637 AAACACAUGUUGCUCAUUGUCUU 6141615-1637 AD-1727700 ACAAUGAGCAACAUGUGUUCU 222 1618-1638AGAACACAUGUUGCUCAUUGUCU 877 1616-1638 AD-1727701 CAAUGAGCAACAUGUGUUCAU223 1619-1639 AUGAACACAUGUUGCUCAUUGUC 878 1617-1639 AD-1727703AUGAGCAACAUGUGUUCAAAU 224 1621-1641 AUUUGAACACAUGUUGCUCAUUG 8791619-1641 AD-1727705 GAGCAACAUGUGUUCAAAGUU 225 1623-1643AACUUUGAACACAUGUUGCUCAU 880 1621-1643 AD-1727708 CAACAUGUGUUCAAAGUCAAU227 1626-1646 AUUGACUUUGAACACAUGUUGCU 881 1624-1646 AD-1727709AACAUGUGUUCAAAGUCAAGU 228 1627-1647 ACUUGACUUUGAACACAUGUUGC 6211625-1647 AD-1727710 ACAUGUGUUCAAAGUCAAGGU 229 1628-1648ACCUUGACUUUGAACACAUGUUG 882 1626-1648 AD-1727712 AUGUGUUCAAAGUCAAGGAUU230 1630-1650 AAUCCUUGACUUUGAACACAUGU 883 1628-1650 AD-1727713UGUGUUCAAAGUCAAGGAUAU 231 1631-1651 AUAUCCUUGACUUUGAACACAUG 8841629-1651 AD-1727714 GUGUUCAAAGUCAAGGAUAUU 232 1632-1652AAUAUCCUUGACUUUGAACACAU 885 1630-1652 AD-1727717 UUCAAAGUCAAGGAUAUGGAU233 1635-1655 AUCCAUAUCCUUGACUUUGAACA 886 1633-1655 AD-1727718UCAAAGUCAAGGAUAUGGAAU 234 1636-1656 AUUCCAUAUCCUUGACUUUGAAC 8871634-1656 AD-1727821 UACCGAUUACCACAAGCAACU 244 1739-1759AGUUGCUUGUGGUAAUCGGUACC 888 1737-1759 AD-1727823 CCGAUUACCACAAGCAACCAU245 1741-1761 AUGGUUGCUUGUGGUAAUCGGUA 889 1739-1761 AD-1727826AUUACCACAAGCAACCAUGGU 247 1744-1764 ACCAUGGUUGCUUGUGGUAAUCG 8901742-1764 AD-1727829 ACCACAAGCAACCAUGGCAGU 250 1747-1767ACUGCCAUGGUUGCUUGUGGUAA 891 1745-1767 AD-1727883 UGGUGUCUGAGUACUUUGUGU259 1822-1842 ACACAAAGUACUCAGACACCACA 892 1820-1842 AD-1727977GAAGCAGGAAUUCCUGAAUUU 263 1974-1994 AAAUUCAGGAAUUCCUGCUUCUU 8931972-1994 AD-1727978 AAGCAGGAAUUCCUGAAUUUU 264 1975-1995AAAAUUCAGGAAUUCCUGCUUCU 894 1973-1995 AD-1727980 GCAGGAAUUCCUGAAUUUUAU265 1977-1997 AUAAAAUUCAGGAAUUCCUGCUU 895 1975-1997 AD-1727981CAGGAAUUCCUGAAUUUUAUU 266 1978-1998 AAUAAAAUUCAGGAAUUCCUGCU 6591976-1998 AD-1727984 GAAUUCCUGAAUUUUAUGACU 269 1981-2001AGUCAUAAAAUUCAGGAAUUCCU 896 1979-2001 AD-1727985 AAUUCCUGAAUUUUAUGACUU270 1982-2002 AAGUCAUAAAAUUCAGGAAUUCC 897 1980-2002 AD-1727986AUUCCUGAAUUUUAUGACUAU 271 1983-2003 AUAGUCAUAAAAUUCAGGAAUUC 8981981-2003 AD-1727987 UUCCUGAAUUUUAUGACUAUU 272 1984-2004AAUAGUCAUAAAAUUCAGGAAUU 665 1982-2004 AD-1727989 CCUGAAUUUUAUGACUAUGAU274 1986-2006 AUCAUAGUCAUAAAAUUCAGGAA 899 1984-2006 AD-1727990CUGAAUUUUAUGACUAUGACU 275 1987-2007 AGUCAUAGUCAUAAAAUUCAGGA 9001985-2007 AD-1727992 GAAUUUUAUGACUAUGACGUU 276 1989-2009AACGUCAUAGUCAUAAAAUUCAG 901 1987-2009 AD-1727993 AAUUUUAUGACUAUGACGUUU277 1990-2010 AAACGUCAUAGUCAUAAAAUUCA 902 1988-2010 AD-1727994AUUUUAUGACUAUGACGUUGU 278 1991-2011 ACAACGUCAUAGUCAUAAAAUUC 9031989-2011 AD-1727996 UUUAUGACUAUGACGUUGCCU 279 1993-2013AGGCAACGUCAUAGUCAUAAAAU 904 1991-2013 AD-1727999 AUGACUAUGACGUUGCCCUGU282 1996-2016 ACAGGGCAACGUCAUAGUCAUAA 905 1994-2016 AD-1728049CAGACUAUCAGGCCCAUUUGU 291 2046-2066 ACAAAUGGGCCUGAUAGUCUGGC 9062044-2066 AD-1728050 AGACUAUCAGGCCCAUUUGUU 292 2047-2067AACAAAUGGGCCUGAUAGUCUGG 907 2045-2067 AD-1728061 CGAGGGAACAACUCGAGCUUU296 2078-2098 AAAGCUCGAGUUGUUCCCUCGGU 908 2076-2098 AD-1728062GAGGGAACAACUCGAGCUUUU 297 2079-2099 AAAAGCUCGAGUUGUUCCCUCGG 9092077-2099 AD-1728067 AACAACUCGAGCUUUGAGGCU 300 2084-2104AGCCUCAAAGCUCGAGUUGUUCC 910 2082-2104 AD-1728085 GCUUCCUCCAACUACCACUUU311 2102-2122 AAAGUGGUAGUUGGAGGAAGCCU 911 2100-2122 AD-1728132CUGCACAGGAUAUCAAAGCUU 319 2149-2169 AAGCUUUGAUAUCCUGUGCAGGG 9122147-2169 AD-1728137 CAGGAUAUCAAAGCUCUGUUU 320 2154-2174AAACAGAGCUUUGAUAUCCUGUG 913 2152-2174 AD-1728140 GAUAUCAAAGCUCUGUUUGUU321 2157-2177 AACAAACAGAGCUUUGAUAUCCU 714 2155-2177 AD-1728146AAAGCUCUGUUUGUGUCUGAU 322 2163-2183 AUCAGACACAAACAGAGCUUUGA 9142161-2183 AD-1728195 AAGAAAGGCAGCUGUGAGAGU 330 2232-2252ACUCUCACAGCUGCCUUUCUUAU 915 2230-2252 AD-1728204 AGCUGUGAGAGAGAUGCUCAU332 2241-2261 AUGAGCAUCUCUCUCACAGCUGC 916 2239-2261 AD-1728206CUGUGAGAGAGAUGCUCAAUU 333 2243-2263 AAUUGAGCAUCUCUCUCACAGCU 9172241-2263 AD-1728207 UGUGAGAGAGAUGCUCAAUAU 334 2244-2264AUAUUGAGCAUCUCUCUCACAGC 918 2242-2264 AD-1728208 GUGAGAGAGAUGCUCAAUAUU335 2245-2265 AAUAUUGAGCAUCUCUCUCACAG 919 2243-2265 AD-1728209UGAGAGAGAUGCUCAAUAUGU 336 2246-2266 ACAUAUUGAGCAUCUCUCUCACA 9202244-2266 AD-1728210 GAGAGAGAUGCUCAAUAUGCU 337 2247-2267AGCAUAUUGAGCAUCUCUCUCAC 921 2245-2267 AD-1728212 CAGGCUAUGACAAAGUCAAGU338 2269-2289 ACUUGACUUUGUCAUAGCCUGGG 922 2267-2289 AD-1728214GGCUAUGACAAAGUCAAGGAU 340 2271-2291 AUCCUUGACUUUGUCAUAGCCUG 9232269-2291 AD-1728220 GACAAAGUCAAGGACAUCUCU 341 2277-2297AGAGAUGUCCUUGACUUUGUCAU 924 2275-2297 AD-1728244 UUGUACUGGAGGAGUGAGUCU347 2321-2341 AGACUCACUCCUCCAGUACAAAG 925 2319-2341 AD-1728258AAUACUUGCAGAGGUGAUUCU 356 2355-2375 AGAAUCACCUCUGCAAGUAUUGG 9262353-2375 AD-1728260 UACUUGCAGAGGUGAUUCUGU 357 2357-2377ACAGAAUCACCUCUGCAAGUAUU 927 2355-2377 AD-1728269 UGAUAGUUCACAAGAGAAGUU360 2386-2406 AACUUCUCUUGUGAACUAUCAAG 928 2384-2406 AD-1728270GAUAGUUCACAAGAGAAGUCU 361 2387-2407 AGACUUCUCUUGUGAACUAUCAA 9292385-2407 AD-1728271 AUAGUUCACAAGAGAAGUCGU 362 2388-2408ACGACUUCUCUUGUGAACUAUCA 930 2386-2408 AD-1728272 UAGUUCACAAGAGAAGUCGUU363 2389-2409 AACGACUUCUCUUGUGAACUAUC 931 2387-2409 AD-1728273AGUUCACAAGAGAAGUCGUUU 364 2390-2410 AAACGACUUCUCUUGUGAACUAU 7572388-2410 AD-1728274 GUUCACAAGAGAAGUCGUUUU 365 2391-2411AAAACGACUUCUCUUGUGAACUA 932 2389-2411 AD-1728275 UUCACAAGAGAAGUCGUUUCU366 2392-2412 AGAAACGACUUCUCUUGUGAACU 759 2390-2412 AD-1728276UCACAAGAGAAGUCGUUUCAU 367 2393-2413 AUGAAACGACUUCUCUUGUGAAC 9332391-2413 AD-1728277 CACAAGAGAAGUCGUUUCAUU 368 2394-2414AAUGAAACGACUUCUCUUGUGAA 934 2392-2414 AD-1728278 ACAAGAGAAGUCGUUUCAUUU369 2395-2415 AAAUGAAACGACUUCUCUUGUGA 762 2393-2415 AD-1728279CAAGAGAAGUCGUUUCAUUCU 370 2396-2416 AGAAUGAAACGACUUCUCUUGUG 9352394-2416 AD-1728280 AAGAGAAGUCGUUUCAUUCAU 371 2397-2417AUGAAUGAAACGACUUCUCUUGU 936 2395-2417 AD-1728282 GAGAAGUCGUUUCAUUCAAGU373 2399-2419 ACUUGAAUGAAACGACUUCUCUU 766 2397-2419 AD-1728283AGAAGUCGUUUCAUUCAAGUU 374 2400-2420 AACUUGAAUGAAACGACUUCUCU 9372398-2420 AD-1728284 GAAGUCGUUUCAUUCAAGUUU 375 2401-2421AAACUUGAAUGAAACGACUUCUC 938 2399-2421 AD-1728285 AAGUCGUUUCAUUCAAGUUGU376 2402-2422 ACAACUUGAAUGAAACGACUUCU 939 2400-2422 AD-1728286AGUCGUUUCAUUCAAGUUGGU 377 2403-2423 ACCAACUUGAAUGAAACGACUUC 9402401-2423 AD-1728300 GAGUAGUGGAUGUCUGCAAAU 378 2437-2457AUUUGCAGACAUCCACUACUCCC 941 2435-2457 AD-1728301 AGUAGUGGAUGUCUGCAAAAU379 2438-2458 AUUUUGCAGACAUCCACUACUCC 942 2436-2458 AD-1728302GUAGUGGAUGUCUGCAAAAAU 380 2439-2459 AUUUUUGCAGACAUCCACUACUC 9432437-2459 AD-1728303 UAGUGGAUGUCUGCAAAAACU 381 2440-2460AGUUUUUGCAGACAUCCACUACU 944 2438-2460 AD-1728307 GGAUGUCUGCAAAAACCAGAU385 2444-2464 AUCUGGUUUUUGCAGACAUCCAC 945 2442-2464 AD-1728308GAUGUCUGCAAAAACCAGAAU 386 2445-2465 AUUCUGGUUUUUGCAGACAUCCA 9462443-2465 AD-1728311 GUCUGCAAAAACCAGAAGCGU 387 2448-2468ACGCUUCUGGUUUUUGCAGACAU 947 2446-2468 AD-1728312 UCUGCAAAAACCAGAAGCGGU388 2449-2469 ACCGCUUCUGGUUUUUGCAGACA 948 2447-2469 AD-1728317AAAAACCAGAAGCGGCAAAAU 391 2454-2474 AUUUUGCCGCUUCUGGUUUUUGC 9492452-2474 AD-1728318 AAAACCAGAAGCGGCAAAAGU 392 2455-2475ACUUUUGCCGCUUCUGGUUUUUG 950 2453-2475 AD-1728320 AACCAGAAGCGGCAAAAGCAU394 2457-2477 AUGCUUUUGCCGCUUCUGGUUUU 951 2455-2477 AD-1728324AGAAGCGGCAAAAGCAGGUAU 395 2461-2481 AUACCUGCUUUUGCCGCUUCUGG 9522459-2481 AD-1728405 AACUCCAAGAUGAGGAUUUGU 399 2542-2562ACAAAUCCUCAUCUUGGAGUUUC 953 2540-2562 AD-1728408 UCCAAGAUGAGGAUUUGGGUU400 2545-2565 AACCCAAAUCCUCAUCUUGGAGU 954 2543-2565 AD-1728410CAAGAUGAGGAUUUGGGUUUU 401 2547-2567 AAAACCCAAAUCCUCAUCUUGGA 7942545-2567 AD-1728412 AGAUGAGGAUUUGGGUUUUCU 402 2549-2569AGAAAACCCAAAUCCUCAUCUUG 795 2547-2569 AD-1728422 GUGGGAUUGAAUUAAAACAGU403 2597-2617 ACUGUUUUAAUUCAAUCCCACGC 955 2595-2617 AD-1728423UGGGAUUGAAUUAAAACAGCU 404 2598-2618 AGCUGUUUUAAUUCAAUCCCACG 9562596-2618 AD-1728424 GGGAUUGAAUUAAAACAGCUU 405 2599-2619AAGCUGUUUUAAUUCAAUCCCAC 957 2597-2619 AD-1728427 AUUGAAUUAAAACAGCUGCGU406 2602-2622 ACGCAGCUGUUUUAAUUCAAUCC 958 2600-2622 AD-1728447AAGGGAAUGUGACCAGGUCUU 19  155-175 AAGACCTGGUCACAUUCCCUUCC 412  153-175AD-1728461 AGGUCUAGGUCUGGAGUUUCU 25  169-189 AGAAACTCCAGACCUAGACCUGG 418 167-189 AD-1728470 UCUGGAGUUUCAGCUUGGACU 27  178-198AGUCCAAGCUGAAACUCCAGACC 420  176-198 AD-1728471 CUGGAGUUUCAGCUUGGACAU 28 179-199 AUGUCCAAGCUGAAACUCCAGAC 959  177-199 AD-1728659UCCUUCUGGCUUCUACCCGUU 31  464-484 AACGGGTAGAAGCCAGAAGGACA 424  462-484AD-1728664 CUGGCUUCUACCCGUACCCUU 34  469-489 AAGGGUACGGGUAGAAGCCAGAA 803 467-489 AD-1728671 CUACCCGUACCCUGUGCAGAU 35  476-496AUCUGCACAGGGUACGGGUAGAA 960  474-496 AD-1728685 UGCAGACACGUACCUGCAGAU 36 490-510 AUCUGCAGGUACGUGUCUGCACA 961  488-510 AD-1728736AAGGCAGAGUGCAGAGCAAUU 37  561-581 AAUUGCTCUGCACUCUGCCUUCC 430  559-581AD-1728777 CCUACUACAAUGUGAGUGAUU 41  634-654 AAUCACTCACAUUGUAGUAGGGA 962 632-654 AD-1728784 CAAUGUGAGUGAUGAGAUCUU 45  641-661AAGATCTCAUCACUCACAUUGUA 438  639-661 AD-1728786 AUGUGAGUGAUGAGAUCUCUU 47 643-663 AAGAGATCUCAUCACUCACAUUG 963  641-663 AD-1728787UGUGAGUGAUGAGAUCUCUUU 48  644-664 AAAGAGAUCUCAUCACUCACAUU 441  642-664AD-1728789 UGAGUGAUGAGAUCUCUUUCU 50  646-666 AGAAAGAGAUCUCAUCACUCACA 808 644-666 AD-1728793 UGAUGAGAUCUCUUUCCACUU 54  650-670AAGUGGAAAGAGAUCUCAUCACU 447  648-670 AD-1728801 UCUCUUUCCACUGCUAUGACU 57 658-678 AGUCAUAGCAGUGGAAAGAGAUC 810  656-678 AD-1728802CUCUUUCCACUGCUAUGACGU 58  659-679 ACGUCATAGCAGUGGAAAGAGAU 451  657-679AD-1728810 ACUGCUAUGACGGUUACACUU 60  667-687 AAGUGUAACCGUCAUAGCAGUGG 811 665-687 AD-1728811 CUGCUAUGACGGUUACACUCU 61  668-688AGAGTGTAACCGUCAUAGCAGUG 454  666-688 AD-1728827 UCGCACCUGCCAAGUGAAUGU 65 704-724 ACAUTCACUUGGCAGGUGCGAUU 458  702-724 AD-1728861CAGACAGCGAUCUGUGACAAU 67  738-758 AUUGTCACAGAUCGCUGUCUGCC 964  736-758AD-1728863 GACAGCGAUCUGUGACAACGU 69  740-760 ACGUTGTCACAGAUCGCUGUCUG 462 738-760 AD-1728877 UGGCACAAGGAAGGUGGGCAU 73  794-814AUGCCCACCUUCCUUGUGCCAAU 965  792-814 AD-1728909 UUGAAGACAGCGUCACCUACU 76 826-846 AGUAGGTGACGCUGUCUUCAAGG 469  824-846 AD-1728990AGACUCCUUCAUGUACGACAU 85  935-955 AUGUCGTACAUGAAGGAGUCUUG 966  933-955AD-1728995 CAAGAGGUGGCCGAAGCUUUU 87  960-980 AAAAGCTUCGGCCACCUCUUGAG 480 958-980 AD-1729031 GAGACCAUAGAAGGAGUCGAU 90  996-1016AUCGACTCCUUCUAUGGUCUCUG 967  994-1016 AD-1729089 CAGGCUCCAUGAACAUCUACU96 1075-1095 AGUAGATGUUCAUGGAGCCUGAA 489 1073-1095 AD-1729103AUCUACCUGGUGCUAGAUGGU 100 1089-1109 ACCATCTAGCACCAGGUAGAUGU 4931087-1109 AD-1729105 CUACCUGGUGCUAGAUGGAUU 102 1091-1111AAUCCATCUAGCACCAGGUAGAU 495 1089-1111 AD-1729106 UACCUGGUGCUAGAUGGAUCU103 1092-1112 AGAUCCAUCUAGCACCAGGUAGA 496 1090-1112 AD-1729110UGGUGCUAGAUGGAUCAGACU 106 1096-1116 AGUCTGAUCCAUCUAGCACCAGG 9681094-1116 AD-1729112 GUGCUAGAUGGAUCAGACAGU 107 1098-1118ACUGTCTGAUCCAUCUAGCACCA 500 1096-1118 AD-1729130 CACAGGAGCCAAAAAGUGUCU116 1136-1156 AGACACTUUUUGGCUCCUGUGAA 509 1134-1156 AD-1729132CAGGAGCCAAAAAGUGUCUAU 118 1138-1158 AUAGACACUUUUUGGCUCCUGUG 9691136-1158 AD-1729134 GGAGCCAAAAAGUGUCUAGUU 120 1140-1160AACUAGACACUUUUUGGCUCCUG 824 1138-1160 AD-1729136 AGCCAAAAAGUGUCUAGUCAU122 1142-1162 AUGACUAGACACUUUUUGGCUCC 970 1140-1162 AD-1729137GCCAAAAAGUGUCUAGUCAAU 123 1143-1163 AUUGACTAGACACUUUUUGGCUC 9711141-1163 AD-1729139 CAAAAAGUGUCUAGUCAACUU 125 1145-1165AAGUTGACUAGACACUUUUUGGC 518 1143-1165 AD-1729141 AAAAGUGUCUAGUCAACUUAU127 1147-1167 AUAAGUTGACUAGACACUUUUUG 972 1145-1167 AD-1729142AAAGUGUCUAGUCAACUUAAU 128 1148-1168 AUUAAGTUGACUAGACACUUUUU 9731146-1168 AD-1729151 AGUCAACUUAAUUGAGAAGGU 134 1157-1177ACCUTCTCAAUUAAGUUGACUAG 974 1155-1177 AD-1729180 AUGGUGUGAAGCCAAGAUAUU143 1186-1206 AAUATCTUGGCUUCACACCAUAA 975 1184-1206 AD-1729207AAAAUUUGGGUCAAAGUGUCU 145 1233-1253 AGACACTUUGACCCAAAUUUUGG 5381231-1253 AD-1729242 UAAUGCAGACUGGGUCACGAU 149 1268-1288AUCGTGACCCAGUCUGCAUUACU 976 1266-1288 AD-1729269 AAUGAAAUCAAUUAUGAAGAU152 1296-1316 AUCUTCAUAAUUGAUUUCAUUGA 977 1294-1316 AD-1729271UGAAAUCAAUUAUGAAGACCU 153 1298-1318 AGGUCUTCAUAAUUGAUUUCAUU 5461296-1318 AD-1729274 AAUCAAUUAUGAAGACCACAU 154 1301-1321AUGUGGTCUUCAUAAUUGAUUUC 978 1299-1321 AD-1729277 CAAUUAUGAAGACCACAAGUU157 1304-1324 AACUTGTGGUCUUCAUAAUUGAU 979 1302-1324 AD-1729280UUAUGAAGACCACAAGUUGAU 160 1307-1327 AUCAACTUGUGGUCUUCAUAAUU 9801305-1327 AD-1729285 AAGACCACAAGUUGAAGUCAU 165 1312-1332AUGACUTCAACUUGUGGUCUUCA 981 1310-1332 AD-1729288 ACCACAAGUUGAAGUCAGGGU167 1315-1335 ACCCTGACUUCAACUUGUGGUCU 560 1313-1335 AD-1729290CACAAGUUGAAGUCAGGGACU 169 1317-1337 AGUCCCTGACUUCAACUUGUGGU 9821315-1337 AD-1729296 UUGAAGUCAGGGACUAACACU 172 1323-1343AGUGTUAGUCCCUGACUUCAACU 565 1321-1343 AD-1729297 UGAAGUCAGGGACUAACACCU173 1324-1344 AGGUGUTAGUCCCUGACUUCAAC 566 1322-1344 AD-1729300AGUCAGGGACUAACACCAAGU 174 1327-1347 ACUUGGTGUUAGUCCCUGACUUC 5671325-1347 AD-1729413 UGAUGGAUUGCACAACAUGGU 181 1442-1462ACCATGTUGUGCAAUCCAUCAGU 574 1440-1462 AD-1729461 UUGGCAAGGAUCGCAAAAACU193 1510-1530 AGUUTUTGCGAUCCUUGCCAAUG 983 1508-1530 AD-1729462UGGCAAGGAUCGCAAAAACCU 194 1511-1531 AGGUTUTUGCGAUCCUUGCCAAU 5871509-1531 AD-1729463 GGCAAGGAUCGCAAAAACCCU 195 1512-1532AGGGTUTUUGCGAUCCUUGCCAA 588 1510-1532 AD-1729487 GAGGAUUAUCUGGAUGUCUAU197 1536-1556 AUAGACAUCCAGAUAAUCCUCCC 861 1534-1556 AD-1729514CCAAGUGAACAUCAAUGCUUU 209 1583-1603 AAAGCATUGAUGUUCACUUGGUU 6021581-1603 AD-1729515 CAAGUGAACAUCAAUGCUUUU 210 1584-1604AAAAGCAUUGAUGUUCACUUGGU 868 1582-1604 AD-1729524 AUCAAUGCUUUGGCUUCCAAU211 1593-1613 AUUGGAAGCCAAAGCAUUGAUGU 869 1591-1613 AD-1729525UCAAUGCUUUGGCUUCCAAGU 212 1594-1614 ACUUGGAAGCCAAAGCAUUGAUG 6051592-1614 AD-1729538 UUCCAAGAAAGACAAUGAGCU 215 1607-1627AGCUCATUGUCUUUCUUGGAAGC 984 1605-1627 AD-1729539 UCCAAGAAAGACAAUGAGCAU216 1608-1628 AUGCTCAUUGUCUUUCUUGGAAG 985 1606-1628 AD-1729541CAAGAAAGACAAUGAGCAACU 217 1610-1630 AGUUGCTCAUUGUCUUUCUUGGA 6101608-1630 AD-1729545 AAAGACAAUGAGCAACAUGUU 219 1614-1634AACATGTUGCUCAUUGUCUUUCU 612 1612-1634 AD-1729548 GACAAUGAGCAACAUGUGUUU221 1617-1637 AAACACAUGUUGCUCAUUGUCUU 614 1615-1637 AD-1729550CAAUGAGCAACAUGUGUUCAU 223 1619-1639 AUGAACACAUGUUGCUCAUUGUC 8781617-1639 AD-1729552 AUGAGCAACAUGUGUUCAAAU 224 1621-1641AUUUGAACACAUGUUGCUCAUUG 879 1619-1641 AD-1729555 AGCAACAUGUGUUCAAAGUCU226 1624-1644 AGACTUTGAACACAUGUUGCUCA 619 1622-1644 AD-1729557CAACAUGUGUUCAAAGUCAAU 227 1626-1646 AUUGACTUUGAACACAUGUUGCU 9861624-1646 AD-1729559 ACAUGUGUUCAAAGUCAAGGU 229 1628-1648ACCUTGACUUUGAACACAUGUUG 622 1626-1648 AD-1729561 AUGUGUUCAAAGUCAAGGAUU230 1630-1650 AAUCCUTGACUUUGAACACAUGU 987 1628-1650 AD-1729562UGUGUUCAAAGUCAAGGAUAU 231 1631-1651 AUAUCCTUGACUUUGAACACAUG 9881629-1651 AD-1729567 UCAAAGUCAAGGAUAUGGAAU 234 1636-1656AUUCCATAUCCUUGACUUUGAAC 989 1634-1656 AD-1729568 CAAAGUCAAGGAUAUGGAAAU235 1637-1657 AUUUCCAUAUCCUUGACUUUGAA 990 1635-1657 AD-1729619UGAAAGCCAGUCUCUGAGUCU 236 1688-1708 AGACTCAGAGACUGGCUUUCAUC 6291686-1708 AD-1729643 UGGCAUGGUUUGGGAACACAU 241 1712-1732AUGUGUTCCCAAACCAUGCCACA 991 1710-1732 AD-1729667 GGGUACCGAUUACCACAAGCU243 1736-1756 AGCUTGTGGUAAUCGGUACCCUU 636 1734-1756 AD-1729670UACCGAUUACCACAAGCAACU 244 1739-1759 AGUUGCTUGUGGUAAUCGGUACC 6371737-1759 AD-1729673 CGAUUACCACAAGCAACCAUU 246 1742-1762AAUGGUTGCUUGUGGUAAUCGGU 639 1740-1762 AD-1729677 UACCACAAGCAACCAUGGCAU249 1746-1766 AUGCCATGGUUGCUUGUGGUAAU 992 1744-1766 AD-1729688ACCAUGGCAGGCCAAGAUCUU 252 1757-1777 AAGATCTUGGCCUGCCAUGGUUG 6451755-1777 AD-1729690 CAUGGCAGGCCAAGAUCUCAU 254 1759-1779AUGAGATCUUGGCCUGCCAUGGU 993 1757-1779 AD-1729729 CUGUGGUGUCUGAGUACUUUU256 1819-1839 AAAAGUACUCAGACACCACAGCC 649 1817-1839 AD-1729802AGCGGGACCUGGAGAUAGAAU 262 1912-1932 AUUCTATCUCCAGGUCCCGCUUC 9941910-1932 AD-1729841 GAAGCAGGAAUUCCUGAAUUU 263 1974-1994AAAUTCAGGAAUUCCUGCUUCUU 995 1972-1994 AD-1729849 AAUUCCUGAAUUUUAUGACUU270 1982-2002 AAGUCATAAAAUUCAGGAAUUCC 996 1980-2002 AD-1729850AUUCCUGAAUUUUAUGACUAU 271 1983-2003 AUAGTCAUAAAAUUCAGGAAUUC 9971981-2003 AD-1729852 UCCUGAAUUUUAUGACUAUGU 273 1985-2005ACAUAGTCAUAAAAUUCAGGAAU 666 1983-2005 AD-1729854 CUGAAUUUUAUGACUAUGACU275 1987-2007 AGUCAUAGUCAUAAAAUUCAGGA 900 1985-2007 AD-1729856GAAUUUUAUGACUAUGACGUU 276 1989-2009 AACGTCAUAGUCAUAAAAUUCAG 6691987-2009 AD-1729861 UUAUGACUAUGACGUUGCCCU 280 1994-2014AGGGCAACGUCAUAGUCAUAAAA 673 1992-2014 AD-1729862 UAUGACUAUGACGUUGCCCUU281 1995-2015 AAGGGCAACGUCAUAGUCAUAAA 674 1993-2015 AD-1729869AUGACGUUGCCCUGAUCAAGU 285 2002-2022 ACUUGATCAGGGCAACGUCAUAG 6782000-2022 AD-1729870 UGACGUUGCCCUGAUCAAGCU 286 2003-2023AGCUTGAUCAGGGCAACGUCAUA 679 2001-2023 AD-1729872 ACGUUGCCCUGAUCAAGCUCU288 2005-2025 AGAGCUTGAUCAGGGCAACGUCA 681 2003-2025 AD-1729926GAGGGAACAACUCGAGCUUUU 297 2079-2099 AAAAGCTCGAGUUGUUCCCUCGG 9982077-2099 AD-1729933 CAACUCGAGCUUUGAGGCUUU 301 2086-2106AAAGCCTCAAAGCUCGAGUUGUU 694 2084-2106 AD-1729941 GCUUUGAGGCUUCCUCCAACU306 2094-2114 AGUUGGAGGAAGCCUCAAAGCUC 699 2092-2114 AD-1729947AGGCUUCCUCCAACUACCACU 310 2100-2120 AGUGGUAGUUGGAGGAAGCCUCA 7032098-2120 AD-1729951 UUCCUCCAACUACCACUUGCU 312 2104-2124AGCAAGTGGUAGUUGGAGGAAGC 705 2102-2124 AD-1729992 CUCCCUGCACAGGAUAUCAAU316 2145-2165 AUUGAUAUCCUGUGCAGGGAGCA 999 2143-2165 AD-1729993UCCCUGCACAGGAUAUCAAAU 317 2146-2166 AUUUGATAUCCUGUGCAGGGAGC 10002144-2166 AD-1729994 CCCUGCACAGGAUAUCAAAGU 318 2147-2167ACUUTGAUAUCCUGUGCAGGGAG 711 2145-2167 AD-1729996 CUGCACAGGAUAUCAAAGCUU319 2149-2169 AAGCTUTGAUAUCCUGUGCAGGG 1001 2147-2169 AD-1730001CAGGAUAUCAAAGCUCUGUUU 320 2154-2174 AAACAGAGCUUUGAUAUCCUGUG 9132152-2174 AD-1730042 GCUGACUCGGAAGGAGGUCUU 323 2195-2215AAGACCTCCUUCCGAGUCAGCUU 716 2193-2215 AD-1730048 UCGGAAGGAGGUCUACAUCAU326 2201-2221 AUGATGTAGACCUCCUUCCGAGU 1002 2199-2221 AD-1730053AGGAGGUCUACAUCAAGAAUU 329 2206-2226 AAUUCUTGAUGUAGACCUCCUUC 10032204-2226 AD-1730059 AAGAAAGGCAGCUGUGAGAGU 330 2232-2252ACUCTCACAGCUGCCUUUCUUAU 1004 2230-2252 AD-1730068 AGCUGUGAGAGAGAUGCUCAU332 2241-2261 AUGAGCAUCUCUCUCACAGCUGC 916 2239-2261 AD-1730071UGUGAGAGAGAUGCUCAAUAU 334 2244-2264 AUAUTGAGCAUCUCUCUCACAGC 10052242-2264 AD-1730077 AGGCUAUGACAAAGUCAAGGU 339 2270-2290ACCUTGACUUUGUCAUAGCCUGG 732 2268-2290 AD-1730103 UUCCUUUGUACUGGAGGAGUU345 2316-2336 AACUCCTCCAGUACAAAGGAACC 1006 2314-2336 AD-1730108UUGUACUGGAGGAGUGAGUCU 347 2321-2341 AGACTCACUCCUCCAGUACAAAG 10072319-2341 AD-1730110 GUACUGGAGGAGUGAGUCCCU 349 2323-2343AGGGACTCACUCCUCCAGUACAA 742 2321-2343 AD-1730112 ACUGGAGGAGUGAGUCCCUAU350 2325-2345 AUAGGGACUCACUCCUCCAGUAC 1008 2323-2345 AD-1730118GGAGUGAGUCCCUAUGCUGAU 353 2331-2351 AUCAGCAUAGGGACUCACUCCUC 10092329-2351 AD-1730122 AAUACUUGCAGAGGUGAUUCU 356 2355-2375AGAATCACCUCUGCAAGUAUUGG 1010 2353-2375 AD-1730133 UGAUAGUUCACAAGAGAAGUU360 2386-2406 AACUTCTCUUGUGAACUAUCAAG 1011 2384-2406 AD-1730143CAAGAGAAGUCGUUUCAUUCU 370 2396-2416 AGAATGAAACGACUUCUCUUGUG 7632394-2416 AD-1730164 GAGUAGUGGAUGUCUGCAAAU 378 2437-2457AUUUGCAGACAUCCACUACUCCC 941 2435-2457 AD-1730167 UAGUGGAUGUCUGCAAAAACU381 2440-2460 AGUUTUTGCAGACAUCCACUACU 774 2438-2460 AD-1730168AGUGGAUGUCUGCAAAAACCU 382 2441-2461 AGGUTUTUGCAGACAUCCACUAC 7752439-2461 AD-1730169 GUGGAUGUCUGCAAAAACCAU 383 2442-2462AUGGTUTUUGCAGACAUCCACUA 1012 2440-2462 AD-1730171 GGAUGUCUGCAAAAACCAGAU385 2444-2464 AUCUGGTUUUUGCAGACAUCCAC 1013 2442-2464 AD-1730183AAACCAGAAGCGGCAAAAGCU 393 2456-2476 AGCUTUTGCCGCUUCUGGUUUUU 7862454-2476 AD-1730184 AACCAGAAGCGGCAAAAGCAU 394 2457-2477AUGCTUTUGCCGCUUCUGGUUUU 1014 2455-2477 AD-1730256 UGGCUGAAGGAGAAACUCCAU398 2529-2549 AUGGAGTUUCUCCUUCAGCCAGG 1015 2527-2549 AD-1730287UGGGAUUGAAUUAAAACAGCU 404 2598-2618 AGCUGUTUUAAUUCAAUCCCACG 10162596-2618 AD-1730288 GGGAUUGAAUUAAAACAGCUU 405 2599-2619AAGCTGTUUUAAUUCAAUCCCAC 798 2597-2619 AD-1730293 UGAAUUAAAACAGCUGCGACU408 2604-2624 AGUCGCAGCUGUUUUAAUUCAAU 1017 2602-2624 AD-1730476AAUUAAAACAGCUGCGACAAU 410 2455-2475 AUUGUCGCAGCUGUUUUAAUUCA 10182453-2475 AD-1730477 AAUUAAAACAGCUGCGACAAU 410 2455-2475ATUGTCGCAGCTGUUUUAAUUCA 1019 2453-2475 AD-1730478 AUUAAAACAGCUGCGACAACU411 2456-2476 AGUUGUCGCAGCUGUUUUAAUUC 1020 2454-2476

TABLE 3Modified Sense and Antisense Strand Sequences of Complement Factor B dsRNA AgentsDuplex Sense Strand Sequence  SEQ ID SEQ ID SEQ Name 5′ to 3′ NO:Antisense Strand Sequence 5′ to 3′ NO: mRNA target sequence NO:AD-1724362 asasgggaauGfUfGfaccaggucuuL96 1021asdAsgadCcdTggucdAcAfuucccuuscsc 1672 GGAAGGGAAUGUGACCAGGUCUA 2385AD-1724363 asgsggaaugUfGfAfccaggucuauL96 1022asdTsagdAcdCuggudCaCfauucccususc 1673 GAAGGGAAUGUGACCAGGUCUAG 2386AD-1724364 gsgsgaauguGfAfCfcaggucuaguL96 1023asdCsuadGadCcuggdTcAfcauucccsusu 1674 AAGGGAAUGUGACCAGGUCUAGG 2387AD-1724365 gsgsaaugugAfCfCfaggucuagguL96 1024asdCscudAgdAccugdGuCfacauuccscsu 1675 AGGGAAUGUGACCAGGUCUAGGU 2388AD-1724369 usgsugaccaGfGfUfcuaggucuguL96 1025asdCsagdAcdCuagadCcUfggucacasusu 1676 AAUGUGACCAGGUCUAGGUCUGG 2389AD-1724370 gsusgaccagGfUfCfuaggucugguL96 1026asdCscadGadCcuagdAcCfuggucacsasu 1677 AUGUGACCAGGUCUAGGUCUGGA 2390AD-1724376 asgsgucuagGfUfCfuggaguuucuL96 1027asdGsaadAcdTccagdAcCfuagaccusgsg 1678 CCAGGUCUAGGUCUGGAGUUUCA 2391AD-1724384 gsuscuggagUfUfUfcagcuuggauL96 1028asdTsccdAadGcugadAaCfuccagacscsu 1679 AGGUCUGGAGUUUCAGCUUGGAC 2392AD-1724385 uscsuggaguUfUfCfagcuuggacuL96 1029asdGsucdCadAgcugdAaAfcuccagascsc 1680 GGUCUGGAGUUUCAGCUUGGACA 2393AD-1724386 csusggaguuUfCfAfgcuuggacauL96 1030asdTsgudCcdAagcudGaAfacuccagsasc 1681 GUCUGGAGUUUCAGCUUGGACAC 2394AD-1724530 uscscuuccgAfCfUfucuccaagauL96 1031asdTscudTgdGagaadGuCfggaaggasgsc 1682 GCUCCUUCCGACUUCUCCAAGAG 2395AD-1724572 usgsuccuucUfGfGfcuucuacccuL96 1032asdGsggdTadGaagcdCaGfaaggacascsa 1683 UGUGUCCUUCUGGCUUCUACCCG 2396AD-1724574 uscscuucugGfCfUfucuacccguuL96 1033asdAscgdGgdTagaadGcCfagaaggascsa 1684 UGUCCUUCUGGCUUCUACCCGUA 2397AD-1724575 cscsuucuggCfUfUfcuacccguauL96 1034asdTsacdGgdGuagadAgCfcagaaggsasc 1685 GUCCUUCUGGCUUCUACCCGUAC 2398AD-1724576 csusucuggcUfUfCfuacceguacuL96 1035asdGsuadCgdGguagdAaGfccagaagsgsa 1686 UCCUUCUGGCUUCUACCCGUACC 2399AD-1724579 csusggcuucUfAfCfccguacccuuL96 1036asdAsggdGudAcgggdTaGfaagccagsasa 1687 UUCUGGCUUCUACCCGUACCCUG 2400AD-1724586 csusacccguAfCfCfcugugcagauL96 1037asdTscudGcdAcaggdGuAfcggguagsasa 1688 UUCUACCCGUACCCUGUGCAGAC 2401AD-1724600 usgscagacaCfGfUfaccugcagauL96 1038asdTscudGcdAgguadCgUfgucugcascsa 1689 UGUGCAGACACGUACCUGCAGAU 2402AD-1724651 asasggcagaGfUfGfcagagcaauuL96 1039asdAsuudGcdTcugcdAcUfcugccuuscsc 1690 GGAAGGCAGAGUGCAGAGCAAUC 2403AD-1724653 gsgscagaguGfCfAfgagcaauccuL96 1040asdGsgadTudGcucudGcAfcucugccsusu 1691 AAGGCAGAGUGCAGAGCAAUCCA 2404AD-1724685 csgsgucuccCfUfAfcuacaauguuL96 1041asdAscadTudGuagudAgGfgagaccgsgsg 1692 CCCGGUCUCCCUACUACAAUGUG 2405AD-1724691 cscscuacuaCfAfAfugugagugauL96 1042asdTscadCudCacaudTgUfaguagggsasg 1693 CUCCCUACUACAAUGUGAGUGAU 2406AD-1724692 cscsuacuacAfAfUfgugagugauuL96 1043asdAsucdAcdTcacadTuGfuaguaggsgsa 1694 UCCCUACUACAAUGUGAGUGAUG 2407AD-1724693 csusacuacaAfUfGfugagugauguL96 1044asdCsaudCadCucacdAuUfguaguagsgsg 1695 CCCUACUACAAUGUGAGUGAUGA 2408AD-1724695 ascsuacaauGfUfGfagugaugaguL96 1045asdCsucdAudCacucdAcAfuuguagusasg 1696 CUACUACAAUGUGAGUGAUGAGA 2409AD-1724698 ascsaaugugAfGfUfgaugagaucuL96 1046asdGsaudCudCaucadCuCfacauugusasg 1697 CUACAAUGUGAGUGAUGAGAUCU 2410AD-1724699 csasaugugaGfUfGfaugagaucuuL96 1047asdAsgadTcdTcaucdAcUfcacauugsusa 1698 UACAAUGUGAGUGAUGAGAUCUC 2411AD-1724700 asasugugagUfGfAfugagaucucuL96 1048asdGsagdAudCucaudCaCfucacauusgsu 1699 ACAAUGUGAGUGAUGAGAUCUCU 2412AD-1724701 asusgugaguGfAfUfgagaucucuuL96 1049asdAsgadGadTcucadTcAfcucacaususg 1700 CAAUGUGAGUGAUGAGAUCUCUU 2413AD-1724702 usgsugagugAfUfGfagaucucuuuL96 1050asdAsagdAgdAucucdAuCfacucacasusu 1701 AAUGUGAGUGAUGAGAUCUCUUU 2414AD-1724703 gsusgagugaUfGfAfgaucucuuuuL96 1051asdAsaadGadGaucudCaUfcacucacsasu 1702 AUGUGAGUGAUGAGAUCUCUUUC 2415AD-1724704 usgsagugauGfAfGfaucucuuucuL96 1052asdGsaadAgdAgaucdTcAfucacucascsa 1703 UGUGAGUGAUGAGAUCUCUUUCC 2416AD-1724705 gsasgugaugAfGfAfucucuuuccuL96 1053asdGsgadAadGagaudCuCfaucacucsasc 1704 GUGAGUGAUGAGAUCUCUUUCCA 2417AD-1724706 asgsugaugaGfAfUfcucuuuccauL96 1054asdTsggdAadAgagadTcUfcaucacuscsa 1705 UGAGUGAUGAGAUCUCUUUCCAC 2418AD-1724707 gsusgaugagAfUfCfucuuuccacuL96 1055asdGsugdGadAagagdAuCfucaucacsusc 1706 GAGUGAUGAGAUCUCUUUCCACU 2419AD-1724708 usgsaugagaUfCfUfcuuuccacuuL96 1056asdAsgudGgdAaagadGaUfcucaucascsu 1707 AGUGAUGAGAUCUCUUUCCACUG 2420AD-1724714 gsasucucuuUfCfCfacugcuauguL96 1057asdCsaudAgdCagugdGaAfagagaucsusc 1708 GAGAUCUCUUUCCACUGCUAUGA 2421AD-1724715 asuscucuuuCfCfAfcugcuaugauL96 1058asdTscadTadGcagudGgAfaagagauscsu 1709 AGAUCUCUUUCCACUGCUAUGAC 2422AD-1724716 uscsucuuucCfAfCfugcuaugacuL96 1059asdGsucdAudAgcagdTgGfaaagagasusc 1710 GAUCUCUUUCCACUGCUAUGACG 2423AD-1724717 csuscuuuccAfCfUfgcuaugacguL96 1060asdCsgudCadTagcadGuGfgaaagagsasu 1711 AUCUCUUUCCACUGCUAUGACGG 2424AD-1724718 uscsuuuccaCfUfGfcuaugacgguL96 1061asdCscgdTcdAuagcdAgUfggaaagasgsa 1712 UCUCUUUCCACUGCUAUGACGGU 2425AD-1724725 ascsugcuauGfAfCfgguuacacuuL96 1062asdAsgudGudAaccgdTcAfuagcagusgsg 1713 CCACUGCUAUGACGGUUACACUC 2426AD-1724726 csusgcuaugAfCfGfguuacacucuL96 1063asdGsagdTgdTaaccdGuCfauagcagsusg 1714 CACUGCUAUGACGGUUACACUCU 2427AD-1724730 usasugacggUfUfAfcacucuccguL96 1064asdCsggdAgdAgugudAaCfcgucauasgsc 1715 GCUAUGACGGUUACACUCUCCGG 2428AD-1724731 asusgacgguUfAfCfacucuccgguL96 1065asdCscgdGadGagugdTaAfccgucausasg 1716 CUAUGACGGUUACACUCUCCGGG 2429AD-1724741 asuscgcaccUfGfCfcaagugaauuL96 1066asdAsuudCadCuuggdCaGfgugcgaususg 1717 CAAUCGCACCUGCCAAGUGAAUG 2430AD-1724742 uscsgcaccuGfCfCfaagugaauguL96 1067asdCsaudTcdAcuugdGcAfggugcgasusu 1718 AAUCGCACCUGCCAAGUGAAUGG 2431AD-1724743 csgscaccugCfCfAfagugaaugguL96 1068asdCscadTudCacuudGgCfaggugcgsasu 1719 AUCGCACCUGCCAAGUGAAUGGC 2432AD-1724776 csasgacagcGfAfUfcugugacaauL96 1069asdTsugdTcdAcagadTcGfcugucugscsc 1720 GGCAGACAGCGAUCUGUGACAAC 2433AD-1724777 asgsacagcgAfUfCfugugacaacuL96 1070asdGsuudGudCacagdAuCfgcugucusgsc 1721 GCAGACAGCGAUCUGUGACAACG 2434AD-1724778 gsascagcgaUfCfUfgugacaacguL96 1071asdCsgudTgdTcacadGaUfcgcugucsusg 1722 CAGACAGCGAUCUGUGACAACGG 2435AD-1724779 ascsagcgauCfUfGfugacaacgguL96 1072asdCscgdTudGucacdAgAfucgcuguscsu 1723 AGACAGCGAUCUGUGACAACGGA 2436AD-1724780 csasgcgaucUfGfUfgacaacggauL96 1073asdTsccdGudTgucadCaGfaucgcugsusc 1724 GACAGCGAUCUGUGACAACGGAG 2437AD-1724781 asgscgaucuGfUfGfacaacggaguL96 1074asdCsucdCgdTugucdAcAfgaucgcusgsu 1725 ACAGCGAUCUGUGACAACGGAGC 2438AD-1724792 usgsgcacaaGfGfAfaggugggcauL96 1075asdTsgcdCcdAccuudCcUfugugccasasu 1726 AUUGGCACAAGGAAGGUGGGCAG 2439AD-1724819 cscsgccuugAfAfGfacagcgucauL96 1076asdTsgadCgdCugucdTuCfaaggcggsusa 1727 UACCGCCUUGAAGACAGCGUCAC 2440AD-1724823 csusugaagaCfAfGfcgucaccuauL96 1077asdTsagdGudGacgcdTgUfcuucaagsgsc 1728 GCCUUGAAGACAGCGUCACCUAC 2441AD-1724824 ususgaagacAfGfCfgucaccuacuL96 1078asdGsuadGgdTgacgdCuGfucuucaasgsg 1729 CCUUGAAGACAGCGUCACCUACC 2442AD-1724825 usgsaagacaGfCfGfucaccuaccuL96 1079asdGsgudAgdGugacdGcUfgucuucasasg 1730 CUUGAAGACAGCGUCACCUACCA 2443AD-1724860 gsusgucaggAfAfGfguggcucuuuL96 1080asdAsagdAgdCcaccdTuCfcugacacsgsu 1731 ACGUGUCAGGAAGGUGGCUCUUG 2444AD-1724894 cscsuuccugCfCfAfagacuccuuuL96 1081asdAsagdGadGucuudGgCfaggaaggscsu 1732 AGCCUUCCUGCCAAGACUCCUUC 2445AD-1724897 uscscugccaAfGfAfcuccuucauuL96 1082asdAsugdAadGgagudCuUfggcaggasasg 1733 CUUCCUGCCAAGACUCCUUCAUG 2446AD-1724899 csusgccaagAfCfUfccuucauguuL96 1083asdAscadTgdAaggadGuCfuuggcagsgsa 1734 UCCUGCCAAGACUCCUUCAUGUA 2447AD-1724900 usgsccaagaCfUfCfcuucauguauL96 1084asdTsacdAudGaaggdAgUfcuuggcasgsg 1735 CCUGCCAAGACUCCUUCAUGUAC 2448AD-1724903 csasagacucCfUfUfcauguacgauL96 1085asdTscgdTadCaugadAgGfagucuugsgsc 1736 GCCAAGACUCCUUCAUGUACGAC 2449AD-1724904 asasgacuccUfUfCfauguacgacuL96 1086asdGsucdGudAcaugdAaGfgagucuusgsg 1737 CCAAGACUCCUUCAUGUACGACA 2450AD-1724905 asgsacuccuUfCfAfuguacgacauL96 1087asdTsgudCgdTacaudGaAfggagucususg 1738 CAAGACUCCUUCAUGUACGACAC 2451AD-1724906 gsascuccuuCfAfUfguacgacacuL96 1088asdGsugdTcdGuacadTgAfaggagucsusu 1739 AAGACUCCUUCAUGUACGACACC 2452AD-1724910 csasagagguGfGfCfcgaagcuuuuL96 1089asdAsaadGcdTucggdCcAfccucuugsasg 1740 CUCAAGAGGUGGCCGAAGCUUUC 2453AD-1724919 gscscgaagcUfUfUfccugucuucuL96 1090asdGsaadGadCaggadAaGfcuucggcscsa 1741 UGGCCGAAGCUUUCCUGUCUUCC 2454AD-1724945 asgsagaccaUfAfGfaaggagucguL96 1091asdCsgadCudCcuucdTaUfggucucusgsu 1742 ACAGAGACCAUAGAAGGAGUCGA 2455AD-1724946 gsasgaccauAfGfAfaggagucgauL96 1092asdTscgdAcdTccuudCuAfuggucucsusg 1743 CAGAGACCAUAGAAGGAGUCGAU 2456AD-1724947 asgsaccauaGfAfAfggagucgauuL96 1093asdAsucdGadCuccudTcUfauggucuscsu 1744 AGAGACCAUAGAAGGAGUCGAUG 2457AD-1724948 gsasccauagAfAfGfgagucgauguL96 1094asdCsaudCgdAcuccdTuCfuauggucsusc 1745 GAGACCAUAGAAGGAGUCGAUGC 2458AD-1724949 ascscauagaAfGfGfagucgaugcuL96 1095asdGscadTedGacucdCuUfcuaugguscsu 1746 AGACCAUAGAAGGAGUCGAUGCU 2459AD-1725000 cscsuucaggCfUfCfcaugaacauuL96 1096asdAsugdTudCauggdAgCfcugaaggsgsu 1747 ACCCUUCAGGCUCCAUGAACAUC 2460AD-1725003 uscsaggcucCfAfUfgaacaucuauL96 1097asdTsagdAudGuucadTgGfagccugasasg 1748 CUUCAGGCUCCAUGAACAUCUAC 2461AD-1725004 csasggcuccAfUfGfaacaucuacuL96 1098asdGsuadGadTguucdAuGfgagccugsasa 1749 UUCAGGCUCCAUGAACAUCUACC 2462AD-1725013 usgsaacaucUfAfCfcuggugcuauL96 1099asdTsagdCadCcaggdTaGfauguucasusg 1750 CAUGAACAUCUACCUGGUGCUAG 2463AD-1725015 asascaucuaCfCfUfggugcuagauL96 1100asdTscudAgdCaccadGgUfagauguuscsa 1751 UGAACAUCUACCUGGUGCUAGAU 2464AD-1725017 csasucuaccUfGfGfugcuagauguL96 1101asdCsaudCudAgcacdCaGfguagaugsusu 1752 AACAUCUACCUGGUGCUAGAUGG 2465AD-1725018 asuscuaccuGfGfUfgcuagaugguL96 1102asdCscadTcdTagcadCcAfgguagausgsu 1753 ACAUCUACCUGGUGCUAGAUGGA 2466AD-1725019 uscsuaccugGfUfGfcuagauggauL96 1103asdTsccdAudCuagcdAcCfagguagasusg 1754 CAUCUACCUGGUGCUAGAUGGAU 2467AD-1725020 csusaccuggUfGfCfuagauggauuL96 1104asdAsucdCadTcuagdCaCfcagguagsasu 1755 AUCUACCUGGUGCUAGAUGGAUC 2468AD-1725021 usasccugguGfCfUfagauggaucuL96 1105asdGsaudCcdAucuadGcAfccagguasgsa 1756 UCUACCUGGUGCUAGAUGGAUCA 2469AD-1725022 ascscuggugCfUfAfgauggaucauL96 1106asdTsgadTcdCaucudAgCfaccaggusasg 1757 CUACCUGGUGCUAGAUGGAUCAG 2470AD-1725023 cscsuggugcUfAfGfauggaucaguL96 1107asdCsugdAudCcaucdTaGfcaccaggsusa 1758 UACCUGGUGCUAGAUGGAUCAGA 2471AD-1725025 usgsgugcuaGfAfUfggaucagacuL96 1108asdGsucdTgdAuccadTcUfagcaccasgsg 1759 CCUGGUGCUAGAUGGAUCAGACA 2472AD-1725027 gsusgcuagaUfGfGfaucagacaguL96 1109asdCsugdTcdTgaucdCaUfcuagcacscsa 1760 UGGUGCUAGAUGGAUCAGACAGC 2473AD-1725028 usgscuagauGfGfAfucagacagcuL96 1110asdGscudGudCugaudCcAfucuagcascsc 1761 GGUGCUAGAUGGAUCAGACAGCA 2474AD-1725033 gsasuggaucAfGfAfcagcauugguL96 1111asdCscadAudGcugudCuGfauccaucsusa 1762 UAGAUGGAUCAGACAGCAUUGGG 2475AD-1725039 csasacuucaCfAfGfgagccaaaauL96 1112asdTsuudTgdGcuccdTgUfgaaguugscsu 1763 AGCAACUUCACAGGAGCCAAAAA 2476AD-1725040 asascuucacAfGfGfagccaaaaauL96 1113asdTsuudTudGgcucdCuGfugaaguusgsc 1764 GCAACUUCACAGGAGCCAAAAAG 2477AD-1725041 ascsuucacaGfGfAfgccaaaaaguL96 1114asdCsuudTudTggcudCcUfgugaagususg 1765 CAACUUCACAGGAGCCAAAAAGU 2478AD-1725042 csusucacagGfAfGfccaaaaaguuL96 1115asdAscudTudTuggcdTcCfugugaagsusu 1766 AACUUCACAGGAGCCAAAAAGUG 2479AD-1725043 ususcacaggAfGfCfcaaaaaguguL96 1116asdCsacdTudTuuggdCuCfcugugaasgsu 1767 ACUUCACAGGAGCCAAAAAGUGU 2480AD-1725044 uscsacaggaGfCfCfaaaaaguguuL96 1117asdAscadCudTuuugdGcUfccugugasasg 1768 CUUCACAGGAGCCAAAAAGUGUC 2481AD-1725045 csascaggagCfCfAfaaaagugucuL96 1118asdGsacdAcdTuuuudGgCfuccugugsasa 1769 UUCACAGGAGCCAAAAAGUGUCU 2482AD-1725046 ascsaggagcCfAfAfaaagugucuuL96 1119asdAsgadCadCuuuudTgGfcuccugusgsa 1770 UCACAGGAGCCAAAAAGUGUCUA 2483AD-1725047 csasggagccAfAfAfaagugucuauL96 1120asdTsagdAcdAcuuudTuGfgcuccugsusg 1771 CACAGGAGCCAAAAAGUGUCUAG 2484AD-1725048 asgsgagccaAfAfAfagugucuaguL96 1121asdCsuadGadCacuudTuUfggcuccusgsu 1772 ACAGGAGCCAAAAAGUGUCUAGU 2485AD-1725049 gsgsagccaaAfAfAfgugucuaguuL96 1122asdAscudAgdAcacudTuUfuggcuccsusg 1773 CAGGAGCCAAAAAGUGUCUAGUC 2486AD-1725050 gsasgccaaaAfAfGfugucuagucuL96 1123asdGsacdTadGacacdTuUfuuggcucscsu 1774 AGGAGCCAAAAAGUGUCUAGUCA 2487AD-1725051 asgsccaaaaAfGfUfgucuagucauL96 1124asdTsgadCudAgacadCuUfuuuggcuscsc 1775 GGAGCCAAAAAGUGUCUAGUCAA 2488AD-1725052 gscscaaaaaGfUfGfucuagucaauL96 1125asdTsugdAcdTagacdAcUfuuuuggcsusc 1776 GAGCCAAAAAGUGUCUAGUCAAC 2489AD-1725053 cscsaaaaagUfGfUfcuagucaacuL96 1126asdGsuudGadCuagadCaCfuuuuuggscsu 1777 AGCCAAAAAGUGUCUAGUCAACU 2490AD-1725054 csasaaaaguGfUfCfuagucaacuuL96 1127asdAsgudTgdAcuagdAcAfcuuuuugsgsc 1778 GCCAAAAAGUGUCUAGUCAACUU 2491AD-1725055 asasaaagugUfCfUfagucaacuuuL96 1128asdAsagdTudGacuadGaCfacuuuuusgsg 1779 CCAAAAAGUGUCUAGUCAACUUA 2492AD-1725056 asasaaguguCfUfAfgucaacuuauL96 1129asdTsaadGudTgacudAgAfcacuuuususg 1780 CAAAAAGUGUCUAGUCAACUUAA 2493AD-1725057 asasagugucUfAfGfucaacuuaauL96 1130asdTsuadAgdTugacdTaGfacacuuususu 1781 AAAAAGUGUCUAGUCAACUUAAU 2494AD-1725058 asasgugucuAfGfUfcaacuuaauuL96 1131asdAsuudAadGuugadCuAfgacacuususu 1782 AAAAGUGUCUAGUCAACUUAAUU 2495AD-1725059 asgsugucuaGfUfCfaacuuaauuuL96 1132asdAsaudTadAguugdAcUfagacacususu 1783 AAAGUGUCUAGUCAACUUAAUUG 2496AD-1725060 gsusgucuagUfCfAfacuuaauuguL96 1133asdCsaadTudAaguudGaCfuagacacsusu 1784 AAGUGUCUAGUCAACUUAAUUGA 2497AD-1725061 usgsucuaguCfAfAfcuuaauugauL96 1134asdTscadAudTaagudTgAfcuagacascsu 1785 AGUGUCUAGUCAACUUAAUUGAG 2498AD-1725062 gsuscuagucAfAfCfuuaauugaguL96 1135asdCsucdAadTuaagdTuGfacuagacsasc 1786 GUGUCUAGUCAACUUAAUUGAGA 2499AD-1725066 asgsucaacuUfAfAfuugagaagguL96 1136asdCscudTcdTcaaudTaAfguugacusasg 1787 CUAGUCAACUUAAUUGAGAAGGU 2500AD-1725074 usasauugagAfAfGfguggcaaguuL96 1137asdAscudTgdCcaccdTuCfucaauuasasg 1788 CUUAAUUGAGAAGGUGGCAAGUU 2501AD-1725075 asasuugagaAfGfGfuggcaaguuuL96 1138asdAsacdTudGccacdCuUfcucaauusasa 1789 UUAAUUGAGAAGGUGGCAAGUUA 2502AD-1725079 gsasgaagguGfGfCfaaguuaugguL96 1139asdCscadTadAcuugdCcAfccuucucsasa 1790 UUGAGAAGGUGGCAAGUUAUGGU 2503AD-1725080 asgsaaggugGfCfAfaguuaugguuL96 1140asdAsccdAudAacuudGcCfaccuucuscsa 1791 UGAGAAGGUGGCAAGUUAUGGUG 2504AD-1725082 asasgguggcAfAfGfuuaugguguuL96 1141asdAscadCcdAuaacdTuGfccaccuuscsu 1792 AGAAGGUGGCAAGUUAUGGUGUG 2505AD-1725083 asgsguggcaAfGfUfuaugguguguL96 1142asdCsacdAcdCauaadCuUfgccaccususc 1793 GAAGGUGGCAAGUUAUGGUGUGA 2506AD-1725088 gscsaaguuaUfGfGfugugaagccuL96 1143asdGsgcdTudCacacdCaUfaacuugcscsa 1794 UGGCAAGUUAUGGUGUGAAGCCA 2507AD-1725092 gsusuaugguGfUfGfaagccaagauL96 1144asdTscudTgdGcuucdAcAfccauaacsusu 1795 AAGUUAUGGUGUGAAGCCAAGAU 2508AD-1725095 asusggugugAfAfGfccaagauauuL96 1145asdAsuadTcdTuggcdTuCfacaccausasa 1796 UUAUGGUGUGAAGCCAAGAUAUG 2509AD-1725096 usgsgugugaAfGfCfcaagauauguL96 1146asdCsaudAudCuuggdCuUfcacaccasusa 1797 UAUGGUGUGAAGCCAAGAUAUGG 2510AD-1725122 asasaauuugGfGfUfcaaagugucuL96 1147asdGsacdAcdTuugadCcCfaaauuuusgsg 1798 CCAAAAUUUGGGUCAAAGUGUCU 2511AD-1725123 asasauuuggGfUfCfaaagugucuuL96 1148asdAsgadCadCuuugdAcCfcaaauuususg 1799 CAAAAUUUGGGUCAAAGUGUCUG 2512AD-1725125 asusuuggguCfAfAfagugucugauL96 1149asdTscadGadCacuudTgAfcccaaaususu 1800 AAAUUUGGGUCAAAGUGUCUGAA 2513AD-1725156 gsusaaugcaGfAfCfugggucacguL96 1150asdCsgudGadCccagdTcUfgcauuacsusg 1801 CAGUAAUGCAGACUGGGUCACGA 2514AD-1725157 usasaugcagAfCfUfgggucacgauL96 1151asdTscgdTgdAcccadGuCfugcauuascsu 1802 AGUAAUGCAGACUGGGUCACGAA 2515AD-1725158 asasugcagaCfUfGfggucacgaauL96 1152asdTsucdGudGacccdAgUfcugcauusasc 1803 GUAAUGCAGACUGGGUCACGAAG 2516AD-1725159 asusgcagacUfGfGfgucacgaaguL96 1153asdCsuudCgdTgaccdCaGfucugcaususa 1804 UAAUGCAGACUGGGUCACGAAGC 2517AD-1725184 asasugaaauCfAfAfuuaugaagauL96 1154asdTscudTcdAuaaudTgAfuuucauusgsa 1805 UCAAUGAAAUCAAUUAUGAAGAC 2518AD-1725186 usgsaaaucaAfUfUfaugaagaccuL96 1155asdGsgudCudTcauadAuUfgauuucasusu 1806 AAUGAAAUCAAUUAUGAAGACCA 2519AD-1725189 asasucaauuAfUfGfaagaccacauL96 1156asdTsgudGgdTcuucdAuAfauugauususc 1807 GAAAUCAAUUAUGAAGACCACAA 2520AD-1725190 asuscaauuaUfGfAfagaccacaauL96 1157asdTsugdTgdGucuudCaUfaauugaususu 1808 AAAUCAAUUAUGAAGACCACAAG 2521AD-1725191 uscsaauuauGfAfAfgaccacaaguL96 1158asdCsuudGudGgucudTcAfuaauugasusu 1809 AAUCAAUUAUGAAGACCACAAGU 2522AD-1725192 csasauuaugAfAfGfaccacaaguuL96 1159asdAscudTgdTggucdTuCfauaauugsasu 1810 AUCAAUUAUGAAGACCACAAGUU 2523AD-1725193 asasuuaugaAfGfAfccacaaguuuL96 1160asdAsacdTudGuggudCuUfcauaauusgsa 1811 UCAAUUAUGAAGACCACAAGUUG 2524AD-1725194 asusuaugaaGfAfCfcacaaguuguL96 1161asdCsaadCudTguggdTcUfucauaaususg 1812 CAAUUAUGAAGACCACAAGUUGA 2525AD-1725195 ususaugaagAfCfCfacaaguugauL96 1162asdTscadAcdTugugdGuCfuucauaasusu 1813 AAUUAUGAAGACCACAAGUUGAA 2526AD-1725196 usasugaagaCfCfAfcaaguugaauL96 1163asdTsucdAadCuugudGgUfcuucauasasu 1814 AUUAUGAAGACCACAAGUUGAAG 2527AD-1725197 asusgaagacCfAfCfaaguugaaguL96 1164asdCsuudCadAcuugdTgGfucuucausasa 1815 UUAUGAAGACCACAAGUUGAAGU 2528AD-1725198 usgsaagaccAfCfAfaguugaaguuL96 1165asdAscudTcdAacuudGuGfgucuucasusa 1816 UAUGAAGACCACAAGUUGAAGUC 2529AD-1725199 gsasagaccaCfAfAfguugaagucuL96 1166asdGsacdTudCaacudTgUfggucuucsasu 1817 AUGAAGACCACAAGUUGAAGUCA 2530AD-1725200 asasgaccacAfAfGfuugaagucauL96 1167asdTsgadCudTcaacdTuGfuggucuuscsa 1818 UGAAGACCACAAGUUGAAGUCAG 2531AD-1725201 asgsaccacaAfGfUfugaagucaguL96 1168asdCsugdAcdTucaadCuUfguggucususc 1819 GAAGACCACAAGUUGAAGUCAGG 2532AD-1725203 ascscacaagUfUfGfaagucaggguL96 1169asdCsccdTgdAcuucdAaCfuugugguscsu 1820 AGACCACAAGUUGAAGUCAGGGA 2533AD-1725204 cscsacaaguUfGfAfagucagggauL96 1170asdTsccdCudGacuudCaAfcuuguggsusc 1821 GACCACAAGUUGAAGUCAGGGAC 2534AD-1725205 csascaaguuGfAfAfgucagggacuL96 1171asdGsucdCcdTgacudTcAfacuugugsgsu 1822 ACCACAAGUUGAAGUCAGGGACU 2535AD-1725206 ascsaaguugAfAfGfucagggacuuL96 1172asdAsgudCcdCugacdTuCfaacuugusgsg 1823 CCACAAGUUGAAGUCAGGGACUA 2536AD-1725208 asasguugaaGfUfCfagggacuaauL96 1173asdTsuadGudCccugdAcUfucaacuusgsu 1824 ACAAGUUGAAGUCAGGGACUAAC 2537AD-1725211 ususgaagucAfGfGfgacuaacacuL96 1174asdGsugdTudAguccdCuGfacuucaascsu 1825 AGUUGAAGUCAGGGACUAACACC 2538AD-1725212 usgsaagucaGfGfGfacuaacaccuL96 1175asdGsgudGudTagucdCcUfgacuucasasc 1826 GUUGAAGUCAGGGACUAACACCA 2539AD-1725215 asgsucagggAfCfUfaacaccaaguL96 1176asdCsuudGgdTguuadGuCfccugacususc 1827 GAAGUCAGGGACUAACACCAAGA 2540AD-1725216 gsuscagggaCfUfAfacaccaagauL96 1177asdTscudTgdGuguudAgUfcccugacsusu 1828 AAGUCAGGGACUAACACCAAGAA 2541AD-1725243 cscsaggcagUfGfUfacagcaugauL96 1178asdTscadTgdCuguadCaCfugccuggsasg 1829 CUCCAGGCAGUGUACAGCAUGAU 2542AD-1725244 csasggcaguGfUfAfcagcaugauuL96 1179asdAsucdAudGcugudAcAfcugccugsgsa 1830 UCCAGGCAGUGUACAGCAUGAUG 2543AD-1725245 asgsgcagugUfAfCfagcaugauguL96 1180asdCsaudCadTgcugdTaCfacugccusgsg 1831 CCAGGCAGUGUACAGCAUGAUGA 2544AD-1725247 gscsaguguaCfAfGfcaugaugaguL96 1181asdCsucdAudCaugcdTgUfacacugcscsu 1832 AGGCAGUGUACAGCAUGAUGAGC 2545AD-1725327 csusgauggaUfUfGfcacaacauguL96 1182asdCsaudGudTgugcdAaUfccaucagsusc 1833 GACUGAUGGAUUGCACAACAUGG 2546AD-1725328 usgsauggauUfGfCfacaacaugguL96 1183asdCscadTgdTugugdCaAfuccaucasgsu 1834 ACUGAUGGAUUGCACAACAUGGG 2547AD-1725329 gsasuggauuGfCfAfcaacauggguL96 1184asdCsccdAudGuugudGcAfauccaucsasg 1835 CUGAUGGAUUGCACAACAUGGGC 2548AD-1725330 asusggauugCfAfCfaacaugggcuL96 1185asdGsccdCadTguugdTgCfaauccauscsa 1836 UGAUGGAUUGCACAACAUGGGCG 2549AD-1725331 usgsgauugcAfCfAfacaugggcguL96 1186asdCsgcdCcdAuguudGuGfcaauccasusc 1837 GAUGGAUUGCACAACAUGGGCGG 2550AD-1725332 gsgsauugcaCfAfAfcaugggcgguL96 1187asdCscgdCcdCaugudTgUfgcaauccsasu 1838 AUGGAUUGCACAACAUGGGCGGG 2551AD-1725333 gsascccaauUfAfCfugucauugauL96 1188asdTscadAudGacagdTaAfuugggucscsc 1839 GGGACCCAAUUACUGUCAUUGAU 2552AD-1725334 ascsccaauuAfCfUfgucauugauuL96 1189asdAsucdAadTgacadGuAfauuggguscsc 1840 GGACCCAAUUACUGUCAUUGAUG 2553AD-1725336 cscsaauuacUfGfUfcauugaugauL96 1190asdTscadTcdAaugadCaGfuaauuggsgsu 1841 ACCCAAUUACUGUCAUUGAUGAG 2554AD-1725344 usgsucauugAfUfGfagauccggguL96 1191asdCsccdGgdAucucdAuCfaaugacasgsu 1842 ACUGUCAUUGAUGAGAUCCGGGA 2555AD-1725345 gsuscauugaUfGfAfgauccgggauL96 1192asdTsccdCgdGaucudCaUfcaaugacsasg 1843 CUGUCAUUGAUGAGAUCCGGGAC 2556AD-1725347 csasuugaugAfGfAfuccgggacuuL96 1193asdAsgudCcdCggaudCuCfaucaaugsasc 1844 GUCAUUGAUGAGAUCCGGGACUU 2557AD-1725348 asusugaugaGfAfUfccgggacuuuL96 1194asdAsagdTcdCcggadTcUfcaucaausgsa 1845 UCAUUGAUGAGAUCCGGGACUUG 2558AD-1725376 ususggcaagGfAfUfcgcaaaaacuL96 1195asdGsuudTudTgcgadTcCfuugccaasusg 1846 CAUUGGCAAGGAUCGCAAAAACC 2559AD-1725377 usgsgcaaggAfUfCfgcaaaaaccuL96 1196asdGsgudTudTugcgdAuCfcuugccasasu 1847 AUUGGCAAGGAUCGCAAAAACCC 2560AD-1725378 gsgscaaggaUfCfGfcaaaaacccuL96 1197asdGsggdTudTuugcdGaUfccuugccsasa 1848 UUGGCAAGGAUCGCAAAAACCCA 2561AD-1725397 csasagggagGfAfUfuaucuggauuL96 1198asdAsucdCadGauaadTcCfucccuugsgsg 1849 CCCAAGGGAGGAUUAUCUGGAUG 2562AD-1725402 gsasggauuaUfCfUfggaugucuauL96 1199asdTsagdAcdAuccadGaUfaauccucscsc 1850 GGGAGGAUUAUCUGGAUGUCUAU 2563AD-1725403 asgsgauuauCfUfGfgaugucuauuL96 1200asdAsuadGadCauccdAgAfuaauccuscsc 1851 GGAGGAUUAUCUGGAUGUCUAUG 2564AD-1725404 gsgsauuaucUfGfGfaugucuauguL96 1201asdCsaudAgdAcaucdCaGfauaauccsusc 1852 GAGGAUUAUCUGGAUGUCUAUGU 2565AD-1725405 gsasuuaucuGfGfAfugucuauguuL96 1202asdAscadTadGacaudCcAfgauaaucscsu 1853 AGGAUUAUCUGGAUGUCUAUGUG 2566AD-1725406 asusuaucugGfAfUfgucuauguguL96 1203asdCsacdAudAgacadTcCfagauaauscsc 1854 GGAUUAUCUGGAUGUCUAUGUGU 2567AD-1725407 ususaucuggAfUfGfucuauguguuL96 1204asdAscadCadTagacdAuCfcagauaasusc 1855 GAUUAUCUGGAUGUCUAUGUGUU 2568AD-1725408 usasucuggaUfGfUfcuauguguuuL96 1205asdAsacdAcdAuagadCaUfccagauasasu 1856 AUUAUCUGGAUGUCUAUGUGUUU 2569AD-1725409 asuscuggauGfUfCfuauguguuuuL96 1206asdAsaadCadCauagdAcAfuccagausasa 1857 UUAUCUGGAUGUCUAUGUGUUUG 2570AD-1725410 uscsuggaugUfCfUfauguguuuguL96 1207asdCsaadAcdAcauadGaCfauccagasusa 1858 UAUCUGGAUGUCUAUGUGUUUGG 2571AD-1725411 csusggauguCfUfAfuguguuugguL96 1208asdCscadAadCacaudAgAfcauccagsasu 1859 AUCUGGAUGUCUAUGUGUUUGGG 2572AD-1725427 asasccaaguGfAfAfcaucaaugcuL96 1209asdGscadTudGaugudTcAfcuugguuscsa 1860 UGAACCAAGUGAACAUCAAUGCU 2573AD-1725428 ascscaagugAfAfCfaucaaugcuuL96 1210asdAsgcdAudTgaugdTuCfacuuggususc 1861 GAACCAAGUGAACAUCAAUGCUU 2574AD-1725429 cscsaagugaAfCfAfucaaugcuuuL96 1211asdAsagdCadTgaudGuUfcacuuggsusu 1862 AACCAAGUGAACAUCAAUGCUUU 2575AD-1725430 csasagugaaCfAfUfcaaugcuuuuL96 1212asdAsaadGcdAuugadTgUfucacuugsgsu 1863 ACCAAGUGAACAUCAAUGCUUUG 2576AD-1725439 asuscaaugcUfUfUfggcuuccaauL96 1213asdTsugdGadAgccadAaGfcauugausgsu 1864 ACAUCAAUGCUUUGGCUUCCAAG 2577AD-1725440 uscsaaugcuUfUfGfgcuuccaaguL96 1214asdCsuudGgdAagccdAaAfgcauugasusg 1865 CAUCAAUGCUUUGGCUUCCAAGA 2578AD-1725441 csasaugcuuUfGfGfcuuccaagauL96 1215asdTscudTgdGaagcdCaAfagcauugsasu 1866 AUCAAUGCUUUGGCUUCCAAGAA 2579AD-1725449 usgsgcuuccAfAfGfaaagacaauuL96 1216asdAsuudGudCuuucdTuGfgaagccasasa 1867 UUUGGCUUCCAAGAAAGACAAUG 2580AD-1725453 ususccaagaAfAfGfacaaugagcuL96 1217asdGscudCadTugucdTuUfcuuggaasgsc 1868 GCUUCCAAGAAAGACAAUGAGCA 2581AD-1725454 uscscaagaaAfGfAfcaaugagcauL96 1218asdTsgcdTcdAuugudCuUfucuuggasasg 1869 CUUCCAAGAAAGACAAUGAGCAA 2582AD-1725456 csasagaaagAfCfAfaugagcaacuL96 1219asdGsuudGcdTcauudGuCfuuucuugsgsa 1870 UCCAAGAAAGACAAUGAGCAACA 2583AD-1725457 asasgaaagaCfAfAfugagcaacauL96 1220asdTsgudTgdCucaudTgUfcuuucuusgsg 1871 CCAAGAAAGACAAUGAGCAACAU 2584AD-1725460 asasagacaaUfGfAfgcaacauguuL96 1221asdAscadTgdTugcudCaUfugucuuuscsu 1872 AGAAAGACAAUGAGCAACAUGUG 2585AD-1725462 asgsacaaugAfGfCfaacauguguuL96 1222asdAscadCadTguugdCuCfauugucususu 1873 AAAGACAAUGAGCAACAUGUGUU 2586AD-1725463 gsascaaugaGfCfAfacauguguuuL96 1223asdAsacdAcdAuguudGcUfcauugucsusu 1874 AAGACAAUGAGCAACAUGUGUUC 2587AD-1725464 ascsaaugagCfAfAfcauguguucuL96 1224asdGsaadCadCaugudTgCfucauuguscsu 1875 AGACAAUGAGCAACAUGUGUUCA 2588AD-1725465 csasaugagcAfAfCfauguguucauL96 1225asdTsgadAcdAcaugdTuGfcucauugsusc 1876 GACAAUGAGCAACAUGUGUUCAA 2589AD-1725467 asusgagcaaCfAfUfguguucaaauL96 1226asdTsuudGadAcacadTgUfugcucaususg 1877 CAAUGAGCAACAUGUGUUCAAAG 2590AD-1725469 gsasgcaacaUfGfUfguucaaaguuL96 1227asdAscudTudGaacadCaUfguugcucsasu 1878 AUGAGCAACAUGUGUUCAAAGUC 2591AD-1725470 asgscaacauGfUfGfuucaaagucuL96 1228asdGsacdTudTgaacdAcAfuguugcuscsa 1879 UGAGCAACAUGUGUUCAAAGUCA 2592AD-1725472 csasacauguGfUfUfcaaagucaauL96 1229asdTsugdAcdTuugadAcAfcauguugscsu 1880 AGCAACAUGUGUUCAAAGUCAAG 2593AD-1725473 asascaugugUfUfCfaaagucaaguL96 1230asdCsuudGadCuuugdAaCfacauguusgsc 1881 GCAACAUGUGUUCAAAGUCAAGG 2594AD-1725474 ascsauguguUfCfAfaagucaagguL96 1231asdCscudTgdAcuuudGaAfcacaugususg 1882 CAACAUGUGUUCAAAGUCAAGGA 2595AD-1725476 asusguguucAfAfAfgucaaggauuL96 1232asdAsucdCudTgacudTuGfaacacausgsu 1883 ACAUGUGUUCAAAGUCAAGGAUA 2596AD-1725477 usgsuguucaAfAfGfucaaggauauL96 1233asdTsaudCcdTugacdTuUfgaacacasusg 1884 CAUGUGUUCAAAGUCAAGGAUAU 2597AD-1725478 gsusguucaaAfGfUfcaaggauauuL96 1234asdAsuadTcdCuugadCuUfugaacacsasu 1885 AUGUGUUCAAAGUCAAGGAUAUG 2598AD-1725481 ususcaaaguCfAfAfggauauggauL96 1235asdTsccdAudAuccudTgAfcuuugaascsa 1886 UGUUCAAAGUCAAGGAUAUGGAA 2599AD-1725482 uscsaaagucAfAfGfgauauggaauL96 1236asdTsucdCadTauccdTuGfacuuugasasc 1887 GUUCAAAGUCAAGGAUAUGGAAA 2600AD-1725483 csasaagucaAfGfGfauauggaaauL96 1237asdTsuudCcdAuaucdCuUfgacuuugsasa 1888 UUCAAAGUCAAGGAUAUGGAAAA 2601AD-1725534 usgsaaagccAfGfUfcucugagucuL96 1238asdGsacdTcdAgagadCuGfgcuuucasusc 1889 GAUGAAAGCCAGUCUCUGAGUCU 2602AD-1725535 gsasaagccaGfUfCfucugagucuuL96 1239asdAsgadCudCagagdAcUfggcuuucsasu 1890 AUGAAAGCCAGUCUCUGAGUCUC 2603AD-1725548 usgsagucucUfGfUfggcaugguuuL96 1240asdAsacdCadTgccadCaGfagacucasgsa 1891 UCUGAGUCUCUGUGGCAUGGUUU 2604AD-1725552 uscsucugugGfCfAfugguuuggguL96 1241asdCsccdAadAccaudGcCfacagagascsu 1892 AGUCUCUGUGGCAUGGUUUGGGA 2605AD-1725556 usgsuggcauGfGfUfuugggaacauL96 1242asdTsgudTcdCcaaadCcAfugccacasgsa 1893 UCUGUGGCAUGGUUUGGGAACAC 2606AD-1725558 usgsgcauggUfUfUfgggaacacauL96 1243asdTsgudGudTcccadAaCfcaugccascsa 1894 UGUGGCAUGGUUUGGGAACACAG 2607AD-1725580 asasggguacCfGfAfuuaccacaauL96 1244asdTsugdTgdGuaaudCgGfuacccuuscsc 1895 GGAAGGGUACCGAUUACCACAAG 2608AD-1725582 gsgsguaccgAfUfUfaccacaagcuL96 1245asdGscudTgdTgguadAuCfgguacccsusu 1896 AAGGGUACCGAUUACCACAAGCA 2609AD-1725585 usasccgauuAfCfCfacaagcaacuL96 1246asdGsuudGcdTugugdGuAfaucgguascsc 1897 GGUACCGAUUACCACAAGCAACC 2610AD-1725587 cscsgauuacCfAfCfaagcaaccauL96 1247asdTsggdTudGcuugdTgGfuaaucggsusa 1898 UACCGAUUACCACAAGCAACCAU 2611AD-1725588 csgsauuaccAfCfAfagcaaccauuL96 1248asdAsugdGudTgcuudGuGfguaaucgsgsu 1899 ACCGAUUACCACAAGCAACCAUG 2612AD-1725590 asusuaccacAfAfGfcaaccaugguL96 1249asdCscadTgdGuugcdTuGfugguaauscsg 1900 CGAUUACCACAAGCAACCAUGGC 2613AD-1725591 ususaccacaAfGfCfaaccauggcuL96 1250asdGsccdAudGguugdCuUfgugguaasusc 1901 GAUUACCACAAGCAACCAUGGCA 2614AD-1725592 usasccacaaGfCfAfaccauggcauL96 1251asdTsgcdCadTgguudGcUfugugguasasu 1902 AUUACCACAAGCAACCAUGGCAG 2615AD-1725593 ascscacaagCfAfAfccauggcaguL96 1252asdCsugdCcdAuggudTgCfuuguggusasa 1903 UUACCACAAGCAACCAUGGCAGG 2616AD-1725598 asasgcaaccAfUfGfgcaggccaauL96 1253asdTsugdGcdCugccdAuGfguugcuusgsu 1904 ACAAGCAACCAUGGCAGGCCAAG 2617AD-1725603 ascscauggcAfGfGfccaagaucuuL96 1254asdAsgadTcdTuggcdCuGfccauggususg 1905 CAACCAUGGCAGGCCAAGAUCUC 2618AD-1725604 cscsauggcaGfGfCfcaagaucucuL96 1255asdGsagdAudCuuggdCcUfgccauggsusu 1906 AACCAUGGCAGGCCAAGAUCUCA 2619AD-1725605 csasuggcagGfCfCfaagaucucauL96 1256asdTsgadGadTcuugdGcCfugccaugsgsu 1907 ACCAUGGCAGGCCAAGAUCUCAG 2620AD-1725643 gscsugugguGfUfCfugaguacuuuL96 1257asdAsagdTadCucagdAcAfccacagescsc 1908 GGGCUGUGGUGUCUGAGUACUUU 2621AD-1725644 csusguggugUfCfUfgaguacuuuuL96 1258asdAsaadGudAcucadGaCfaccacagscsc 1909 GGCUGUGGUGUCUGAGUACUUUG 2622AD-1725645 usgsugguguCfUfGfaguacuuuguL96 1259asdCsaadAgdTacucdAgAfcaccacasgsc 1910 GCUGUGGUGUCUGAGUACUUUGU 2623AD-1725646 gsusggugucUfGfAfguacuuuguuL96 1260asdAscadAadGuacudCaGfacaccacsasg 1911 CUGUGGUGUCUGAGUACUUUGUG 2624AD-1725647 usgsgugucuGfAfGfuacuuuguguL96 1261asdCsacdAadAguacdTcAfgacaccascsa 1912 UGUGGUGUCUGAGUACUUUGUGC 2625AD-1725667 csusgacagcAfGfCfacauuguuuuL96 1262asdAsaadCadAugugdCuGfcugucagscsa 1913 UGCUGACAGCAGCACAUUGUUUC 2626AD-1725716 asasgcgggaCfCfUfggagauagauL96 1263asdTscudAudCuccadGgUfcccgcuuscsu 1914 AGAAGCGGGACCUGGAGAUAGAA 2627AD-1725717 asgscgggacCfUfGfgagauagaauL96 1264asdTsucdTadTcuccdAgGfucccgcususc 1915 GAAGCGGGACCUGGAGAUAGAAG 2628AD-1725756 gsasagcaggAfAfUfuccugaauuuL96 1265asdAsaudTcdAggaadTuCfcugcuucsusu 1916 AAGAAGCAGGAAUUCCUGAAUUU 2629AD-1725757 asasgcaggaAfUfUfccugaauuuuL96 1266asdAsaadTudCaggadAuUfccugcuuscsu 1917 AGAAGCAGGAAUUCCUGAAUUUU 2630AD-1725759 gscsaggaauUfCfCfugaauuuuauL96 1267asdTsaadAadTucagdGaAfuuccugcsusu 1918 AAGCAGGAAUUCCUGAAUUUUAU 2631AD-1725760 csasggaauuCfCfUfgaauuuuauuL96 1268asdAsuadAadAuucadGgAfauuccugscsu 1919 AGCAGGAAUUCCUGAAUUUUAUG 2632AD-1725761 asgsgaauucCfUfGfaauuuuauguL96 1269asdCsaudAadAauucdAgGfaauuccusgsc 1920 GCAGGAAUUCCUGAAUUUUAUGA 2633AD-1725762 gsgsaauuccUfGfAfauuuuaugauL96 1270asdTscadTadAaauudCaGfgaauuccsusg 1921 CAGGAAUUCCUGAAUUUUAUGAC 2634AD-1725763 gsasauuccuGfAfAfuuuuaugacuL96 1271asdGsucdAudAaaaudTcAfggaauucscsu 1922 AGGAAUUCCUGAAUUUUAUGACU 2635AD-1725764 asasuuccugAfAfUfuuuaugacuuL96 1272asdAsgudCadTaaaadTuCfaggaauuscsc 1923 GGAAUUCCUGAAUUUUAUGACUA 2636AD-1725765 asusuccugaAfUfUfuuaugacuauL96 1273asdTsagdTcdAuaaadAuUfcaggaaususc 1924 GAAUUCCUGAAUUUUAUGACUAU 2637AD-1725766 ususccugaaUfUfUfuaugacuauuL96 1274asdAsuadGudCauaadAaUfucaggaasusu 1925 AAUUCCUGAAUUUUAUGACUAUG 2638AD-1725767 uscscugaauUfUfUfaugacuauguL96 1275asdCsaudAgdTcauadAaAfuucaggasasu 1926 AUUCCUGAAUUUUAUGACUAUGA 2639AD-1725768 cscsugaauuUfUfAfugacuaugauL96 1276asdTscadTadGucaudAaAfauucaggsasa 1927 UUCCUGAAUUUUAUGACUAUGAC 2640AD-1725769 csusgaauuuUfAfUfgacuaugacuL96 1277asdGsucdAudAgucadTaAfaauucagsgsa 1928 UCCUGAAUUUUAUGACUAUGACG 2641AD-1725771 gsasauuuuaUfGfAfcuaugacguuL96 1278asdAscgdTcdAuagudCaUfaaaauucsasg 1929 CUGAAUUUUAUGACUAUGACGUU 2642AD-1725772 asasuuuuauGfAfCfuaugacguuuL96 1279asdAsacdGudCauagdTcAfuaaaauuscsa 1930 UGAAUUUUAUGACUAUGACGUUG 2643AD-1725773 asusuuuaugAfCfUfaugacguuguL96 1280asdCsaadCgdTcauadGuCfauaaaaususc 1931 GAAUUUUAUGACUAUGACGUUGC 2644AD-1725775 ususuaugacUfAfUfgacguugccuL96 1281asdGsgcdAadCgucadTaGfucauaaasasu 1932 AUUUUAUGACUAUGACGUUGCCC 2645AD-1725776 ususaugacuAfUfGfacguugcccuL96 1282asdGsggdCadAcgucdAuAfgucauaasasa 1933 UUUUAUGACUAUGACGUUGCCCU 2646AD-1725777 usasugacuaUfGfAfcguugcccuuL96 1283asdAsggdGcdAacgudCaUfagucauasasa 1934 UUUAUGACUAUGACGUUGCCCUG 2647AD-1725778 asusgacuauGfAfCfguugcccuguL96 1284asdCsagdGgdCaacgdTcAfuagucausasa 1935 UUAUGACUAUGACGUUGCCCUGA 2648AD-1725779 usgsacuaugAfCfGfuugcccugauL96 1285asdTscadGgdGcaacdGuCfauagucasusa 1936 UAUGACUAUGACGUUGCCCUGAU 2649AD-1725780 gsascuaugaCfGfUfugcccugauuL96 1286asdAsucdAgdGgcaadCgUfcauagucsasu 1937 AUGACUAUGACGUUGCCCUGAUC 2650AD-1725784 asusgacguuGfCfCfcugaucaaguL96 1287asdCsuudGadTcaggdGcAfacgucausasg 1938 CUAUGACGUUGCCCUGAUCAAGC 2651AD-1725785 usgsacguugCfCfCfugaucaagcuL96 1288asdGscudTgdAucagdGgCfaacgucasusa 1939 UAUGACGUUGCCCUGAUCAAGCU 2652AD-1725786 gsascguugcCfCfUfgaucaagcuuL96 1289asdAsgcdTudGaucadGgGfcaacgucsasu 1940 AUGACGUUGCCCUGAUCAAGCUC 2653AD-1725787 ascsguugccCfUfGfaucaagcucuL96 1290asdGsagdCudTgaucdAgGfgcaacguscsa 1941 UGACGUUGCCCUGAUCAAGCUCA 2654AD-1725789 gsusugcccuGfAfUfcaagcucaauL96 1291asdTsugdAgdCuugadTcAfgggcaacsgsu 1942 ACGUUGCCCUGAUCAAGCUCAAG 2655AD-1725790 ususgcccugAfUfCfaagcucaaguL96 1292asdCsuudGadGcuugdAuCfagggcaascsg 1943 CGUUGCCCUGAUCAAGCUCAAGA 2656AD-1725828 csasgacuauCfAfGfgcccauuuguL96 1293asdCsaadAudGggccdTgAfuagucugsgsc 1944 GCCAGACUAUCAGGCCCAUUUGU 2657AD-1725829 asgsacuaucAfGfGfcccauuuguuL96 1294asdAscadAadTgggcdCuGfauagucusgsg 1945 CCAGACUAUCAGGCCCAUUUGUC 2658AD-1725830 gsascuaucaGfGfCfccauuugucuL96 1295asdGsacdAadAugggdCcUfgauagucsusg 1946 CAGACUAUCAGGCCCAUUUGUCU 2659AD-1725831 ascsuaucagGfCfCfcauuugucuuL96 1296asdAsgadCadAauggdGcCfugauaguscsu 1947 AGACUAUCAGGCCCAUUUGUCUC 2660AD-1725832 csusaucaggCfCfCfauuugucucuL96 1297asdGsagdAcdAaaugdGgCfcugauagsusc 1948 GACUAUCAGGCCCAUUUGUCUCC 2661AD-1725840 csgsagggaaCfAfAfcucgagcuuuL96 1298asdAsagdCudCgagudTgUfucccucgsgsu 1949 ACCGAGGGAACAACUCGAGCUUU 2662AD-1725841 gsasgggaacAfAfCfucgagcuuuuL96 1299asdAsaadGcdTcgagdTuGfuucccucsgsg 1950 CCGAGGGAACAACUCGAGCUUUG 2663AD-1725842 asgsggaacaAfCfUfcgagcuuuguL96 1300asdCsaadAgdCucgadGuUfguucccuscsg 1951 CGAGGGAACAACUCGAGCUUUGA 2664AD-1725845 gsasacaacuCfGfAfgcuuugagguL96 1301asdCscudCadAagcudCgAfguuguucscsc 1952 GGGAACAACUCGAGCUUUGAGGC 2665AD-1725846 asascaacucGfAfGfcuuugaggcuL96 1302asdGsccdTcdAaagcdTcGfaguuguuscsc 1953 GGAACAACUCGAGCUUUGAGGCU 2666AD-1725848 csasacucgaGfCfUfuugaggcuuuL96 1303asdAsagdCcdTcaaadGcUfcgaguugsusu 1954 AACAACUCGAGCUUUGAGGCUUC 2667AD-1725849 asascucgagCfUfUfugaggcuucuL96 1304asdGsaadGcdCucaadAgCfucgaguusgsu 1955 ACAACUCGAGCUUUGAGGCUUCC 2668AD-1725850 ascsucgagcUfUfUfgaggcuuccuL96 1305asdGsgadAgdCcucadAaGfcucgagususg 1956 CAACUCGAGCUUUGAGGCUUCCU 2669AD-1725854 gsasgcuuugAfGfGfcuuccuccauL96 1306asdTsggdAgdGaagcdCuCfaaagcucsgsa 1957 UCGAGCUUUGAGGCUUCCUCCAA 2670AD-1725855 asgscuuugaGfGfCfuuccuccaauL96 1307asdTsugdGadGgaagdCcUfcaaagcuscsg 1958 CGAGCUUUGAGGCUUCCUCCAAC 2671AD-1725856 gscsuuugagGfCfUfuccuccaacuL96 1308asdGsuudGgdAggaadGcCfucaaagcsusc 1959 GAGCUUUGAGGCUUCCUCCAACU 2672AD-1725857 csusuugaggCfUfUfccuccaacuuL96 1309asdAsgudTgdGaggadAgCfcucaaagscsu 1960 AGCUUUGAGGCUUCCUCCAACUA 2673AD-1725858 ususugaggcUfUfCfcuccaacuauL96 1310asdTsagdTudGgaggdAaGfccucaaasgsc 1961 GCUUUGAGGCUUCCUCCAACUAC 2674AD-1725861 gsasggcuucCfUfCfcaacuaccauL96 1311asdTsggdTadGuuggdAgGfaagccucsasa 1962 UUGAGGCUUCCUCCAACUACCAC 2675AD-1725862 asgsgcuuccUfCfCfaacuaccacuL96 1312asdGsugdGudAguugdGaGfgaagccuscsa 1963 UGAGGCUUCCUCCAACUACCACU 2676AD-1725864 gscsuuccucCfAfAfcuaccacuuuL96 1313asdAsagdTgdGuagudTgGfaggaagcscsu 1964 AGGCUUCCUCCAACUACCACUUG 2677AD-1725866 ususccuccaAfCfUfaccacuugcuL96 1314asdGscadAgdTgguadGuUfggaggaasgsc 1965 GCUUCCUCCAACUACCACUUGCC 2678AD-1725867 uscscuccaaCfUfAfccacuugccuL96 1315asdGsgcdAadGuggudAgUfuggaggasasg 1966 CUUCCUCCAACUACCACUUGCCA 2679AD-1725872 csasacuaccAfCfUfugccagcaauL96 1316asdTsugdCudGgcaadGuGfguaguugsgsa 1967 UCCAACUACCACUUGCCAGCAAC 2680AD-1725874 ascsuaccacUfUfGfccagcaacauL96 1317asdTsgudTgdCuggcdAaGfugguagususg 1968 CAACUACCACUUGCCAGCAACAA 2681AD-1725907 csuscccugcAfCfAfggauaucaauL96 1318asdTsugdAudAuccudGuGfcagggagscsa 1969 UGCUCCCUGCACAGGAUAUCAAA 2682AD-1725908 uscsccugcaCfAfGfgauaucaaauL96 1319asdTsuudGadTauccdTgUfgcagggasgsc 1970 GCUCCCUGCACAGGAUAUCAAAG 2683AD-1725909 cscscugcacAfGfGfauaucaaaguL96 1320asdCsuudTgdAuaucdCuGfugcagggsasg 1971 CUCCCUGCACAGGAUAUCAAAGC 2684AD-1725911 csusgcacagGfAfUfaucaaagcuuL96 1321asdAsgcdTudTgauadTcCfugugcagsgsg 1972 CCCUGCACAGGAUAUCAAAGCUC 2685AD-1725916 csasggauauCfAfAfagcucuguuuL96 1322asdAsacdAgdAgcuudTgAfuauccugsusg 1973 CACAGGAUAUCAAAGCUCUGUUU 2686AD-1725919 gsasuaucaaAfGfCfucuguuuguuL96 1323asdAscadAadCagagdCuUfugauaucscsu 1974 AGGAUAUCAAAGCUCUGUUUGUG 2687AD-1725925 asasagcucuGfUfUfugugucugauL96 1324asdTscadGadCacaadAcAfgagcuuusgsa 1975 UCAAAGCUCUGUUUGUGUCUGAG 2688AD-1725957 gscsugacucGfGfAfaggaggucuuL96 1325asdAsgadCcdTccuudCcGfagucagesusu 1976 AAGCUGACUCGGAAGGAGGUCUA 2689AD-1725958 csusgacucgGfAfAfggaggucuauL96 1326asdTsagdAcdCuccudTcCfgagucagscsu 1977 AGCUGACUCGGAAGGAGGUCUAC 2690AD-1725961 ascsucggaaGfGfAfggucuacauuL96 1327asdAsugdTadGaccudCcUfuccgaguscsa 1978 UGACUCGGAAGGAGGUCUACAUC 2691AD-1725963 uscsggaaggAfGfGfucuacaucauL96 1328asdTsgadTgdTagacdCuCfcuuccgasgsu 1979 ACUCGGAAGGAGGUCUACAUCAA 2692AD-1725964 csgsgaaggaGfGfUfcuacaucaauL96 1329asdTsugdAudGuagadCcUfccuuccgsasg 1980 CUCGGAAGGAGGUCUACAUCAAG 2693AD-1725967 asasggagguCfUfAfcaucaagaauL96 1330asdTsucdTudGaugudAgAfccuccuuscsc 1981 GGAAGGAGGUCUACAUCAAGAAU 2694AD-1725968 asgsgaggucUfAfCfaucaagaauuL96 1331asdAsuudCudT gaugdTaGfaccuccususc 1982 GAAGGAGGUCUACAUCAAGAAUG 2695AD-1725974 asasgaaaggCfAfGfcugugagaguL96 1332asdCsucdTcdAcagedTgCfcuuucuusasu 1983 AUAAGAAAGGCAGCUGUGAGAGA 2696AD-1725977 asasaggcagCfUfGfugagagagauL96 1333asdTscudCudCucacdAgCfugccuuuscsu 1984 AGAAAGGCAGCUGUGAGAGAGAU 2697AD-1725983 asgscugugaGfAfGfagaugcucauL96 1334asdTsgadGcdAucucdTcUfcacagcusgsc 1985 GCAGCUGUGAGAGAGAUGCUCAA 2698AD-1725985 csusgugagaGfAfGfaugcucaauuL96 1335asdAsuudGadGcaucdTcUfcucacagscsu 1986 AGCUGUGAGAGAGAUGCUCAAUA 2699AD-1725986 usgsugagagAfGfAfugcucaauauL96 1336asdTsaudTgdAgcaudCuCfucucacasgsc 1987 GCUGUGAGAGAGAUGCUCAAUAU 2700AD-1725987 gsusgagagaGfAfUfgcucaauauuL96 1337asdAsuadTudGagcadTcUfcucucacsasg 1988 CUGUGAGAGAGAUGCUCAAUAUG 2701AD-1725988 usgsagagagAfUfGfcucaauauguL96 1338asdCsaudAudTgagcdAuCfucucucascsa 1989 UGUGAGAGAGAUGCUCAAUAUGC 2702AD-1725989 gsasgagagaUfGfCfucaauaugcuL96 1339asdGscadTadTugagdCaUfcucucucsasc 1990 GUGAGAGAGAUGCUCAAUAUGCC 2703AD-1725991 csasggcuauGfAfCfaaagucaaguL96 1340asdCsuudGadCuuugdTcAfuagccugsgsg 1991 CCCAGGCUAUGACAAAGUCAAGG 2704AD-1725992 asgsgcuaugAfCfAfaagucaagguL96 1341asdCscudTgdAcuuudGuCfauagccusgsg 1992 CCAGGCUAUGACAAAGUCAAGGA 2705AD-1725993 gsgscuaugaCfAfAfagucaaggauL96 1342asdTsccdTudGacuudTgUfcauagccsusg 1993 CAGGCUAUGACAAAGUCAAGGAC 2706AD-1725999 gsascaaaguCfAfAfggacaucucuL96 1343asdGsagdAudGuccudTgAfcuuugucsasu 1994 AUGACAAAGUCAAGGACAUCUCA 2707AD-1726014 uscsgguuccUfUfUfguacuggaguL96 1344asdCsucdCadGuacadAaGfgaaccgasgsg 1995 CCUCGGUUCCUUUGUACUGGAGG 2708AD-1726015 csgsguuccuUfUfGfuacuggagguL96 1345asdCscudCcdAguacdAaAfggaaccgsasg 1996 CUCGGUUCCUUUGUACUGGAGGA 2709AD-1726016 gsgsuuccuuUfGfUfacuggaggauL96 1346asdTsccdTcdCaguadCaAfaggaaccsgsa 1997 UCGGUUCCUUUGUACUGGAGGAG 2710AD-1726018 ususccuuugUfAfCfuggaggaguuL96 1347asdAscudCcdTccagdTaCfaaaggaascsc 1998 GGUUCCUUUGUACUGGAGGAGUG 2711AD-1726020 cscsuuuguaCfUfGfgaggagugauL96 1348asdTscadCudCcuccdAgUfacaaaggsasa 1999 UUCCUUUGUACUGGAGGAGUGAG 2712AD-1726023 ususguacugGfAfGfgagugagucuL96 1349asdGsacdTcdAcuccdTcCfaguacaasasg 2000 CUUUGUACUGGAGGAGUGAGUCC 2713AD-1726024 usgsuacuggAfGfGfagugaguccuL96 1350asdGsgadCudCacucdCuCfcaguacasasa 2001 UUUGUACUGGAGGAGUGAGUCCC 2714AD-1726025 gsusacuggaGfGfAfgugagucccuL96 1351asdGsggdAcdTcacudCcUfccaguacsasa 2002 UUGUACUGGAGGAGUGAGUCCCU 2715AD-1726027 ascsuggaggAfGfUfgagucccuauL96 1352asdTsagdGgdAcucadCuCfcuccagusasc 2003 GUACUGGAGGAGUGAGUCCCUAU 2716AD-1726029 usgsgaggagUfGfAfgucccuauguL96 1353asdCsaudAgdGgacudCaCfuccuccasgsu 2004 ACUGGAGGAGUGAGUCCCUAUGC 2717AD-1726031 gsasggagugAfGfUfcccuaugcuuL96 1354asdAsgcdAudAgggadCuCfacuccucscsa 2005 UGGAGGAGUGAGUCCCUAUGCUG 2718AD-1726033 gsgsagugagUfCfCfcuaugcugauL96 1355asdTscadGcdAuaggdGaCfucacuccsusc 2006 GAGGAGUGAGUCCCUAUGCUGAC 2719AD-1726034 gsasgugaguCfCfCfuaugcugacuL96 1356asdGsucdAgdCauagdGgAfcucacucscsu 2007 AGGAGUGAGUCCCUAUGCUGACC 2720AD-1726036 csasauacuuGfCfAfgaggugauuuL96 1357asdAsaudCadCcucudGcAfaguauugsgsg 2008 CCCAAUACUUGCAGAGGUGAUUC 2721AD-1726037 asasuacuugCfAfGfaggugauucuL96 1358asdGsaadTcdAccucdTgCfaaguauusgsg 2009 CCAAUACUUGCAGAGGUGAUUCU 2722AD-1726039 usascuugcaGfAfGfgugauucuguL96 1359asdCsagdAadTcaccdTcUfgcaaguasusu 2010 AAUACUUGCAGAGGUGAUUCUGG 2723AD-1726041 csusugcagaGfGfUfgauucuggcuL96 1360asdGsccdAgdAaucadCcUfcugcaagsusa 2011 UACUUGCAGAGGUGAUUCUGGCG 2724AD-1726042 ususgcagagGfUfGfauucuggcguL96 1361asdCsgcdCadGaaucdAcCfucugcaasgsu 2012 ACUUGCAGAGGUGAUUCUGGCGG 2725AD-1726048 usgsauaguuCfAfCfaagagaaguuL96 1362asdAscudTcdTcuugdTgAfacuaucasasg 2013 CUUGAUAGUUCACAAGAGAAGUC 2726AD-1726049 gsasuaguucAfCfAfagagaagucuL96 1363asdGsacdTudCucuudGuGfaacuaucsasa 2014 UUGAUAGUUCACAAGAGAAGUCG 2727AD-1726050 asusaguucaCfAfAfgagaagucguL96 1364asdCsgadCudTcucudTgUfgaacuauscsa 2015 UGAUAGUUCACAAGAGAAGUCGU 2728AD-1726051 usasguucacAfAfGfagaagucguuL96 1365asdAscgdAcdTucucdTuGfugaacuasusc 2016 GAUAGUUCACAAGAGAAGUCGUU 2729AD-1726052 asgsuucacaAfGfAfgaagucguuuL96 1366asdAsacdGadCuucudCuUfgugaacusasu 2017 AUAGUUCACAAGAGAAGUCGUUU 2730AD-1726053 gsusucacaaGfAfGfaagucguuuuL96 1367asdAsaadCgdAcuucdTcUfugugaacsusa 2018 UAGUUCACAAGAGAAGUCGUUUC 2731AD-1726054 ususcacaagAfGfAfagucguuucuL96 1368asdGsaadAcdGacuudCuCfuugugaascsu 2019 AGUUCACAAGAGAAGUCGUUUCA 2732AD-1726055 uscsacaagaGfAfAfgucguuucauL96 1369asdTsgadAadCgacudTcUfcuugugasasc 2020 GUUCACAAGAGAAGUCGUUUCAU 2733AD-1726056 csascaagagAfAfGfucguuucauuL96 1370asdAsugdAadAcgacdTuCfucuugugsasa 2021 UUCACAAGAGAAGUCGUUUCAUU 2734AD-1726057 ascsaagagaAfGfUfcguuucauuuL96 1371asdAsaudGadAacgadCuUfcucuugusgsa 2022 UCACAAGAGAAGUCGUUUCAUUC 2735AD-1726058 csasagagaaGfUfCfguuucauucuL96 1372asdGsaadTgdAaacgdAcUfucucuugsusg 2023 CACAAGAGAAGUCGUUUCAUUCA 2736AD-1726059 asasgagaagUfCfGfuuucauucauL96 1373asdTsgadAudGaaacdGaCfuucucuusgsu 2024 ACAAGAGAAGUCGUUUCAUUCAA 2737AD-1726060 asgsagaaguCfGfUfuucauucaauL96 1374asdTsugdAadTgaaadCgAfcuucucususg 2025 CAAGAGAAGUCGUUUCAUUCAAG 2738AD-1726061 gsasgaagucGfUfUfucauucaaguL96 1375asdCsuudGadAugaadAcGfacuucucsusu 2026 AAGAGAAGUCGUUUCAUUCAAGU 2739AD-1726062 asgsaagucgUfUfUfcauucaaguuL96 1376asdAscudTgdAaugadAaCfgacuucuscsu 2027 AGAGAAGUCGUUUCAUUCAAGUU 2740AD-1726063 gsasagucguUfUfCfauucaaguuuL96 1377asdAsacdTudGaaugdAaAfcgacuucsusc 2028 GAGAAGUCGUUUCAUUCAAGUUG 2741AD-1726064 asasgucguuUfCfAfuucaaguuguL96 1378asdCsaadCudTgaaudGaAfacgacuuscsu 2029 AGAAGUCGUUUCAUUCAAGUUGG 2742AD-1726065 asgsucguuuCfAfUfucaaguugguL96 1379asdCscadAcdTugaadTgAfaacgacususc 2030 GAAGUCGUUUCAUUCAAGUUGGU 2743AD-1726079 gsasguagugGfAfUfgucugcaaauL96 1380asdTsuudGcdAgacadTcCfacuacucscsc 2031 GGGAGUAGUGGAUGUCUGCAAAA 2744AD-1726080 asgsuaguggAfUfGfucugcaaaauL96 1381asdTsuudTgdCagacdAuCfcacuacuscsc 2032 GGAGUAGUGGAUGUCUGCAAAAA 2745AD-1726081 gsusaguggaUfGfUfcugcaaaaauL96 1382asdTsuudTudGcagadCaUfccacuacsusc 2033 GAGUAGUGGAUGUCUGCAAAAAC 2746AD-1726082 usasguggauGfUfCfugcaaaaacuL96 1383asdGsuudTudTgcagdAcAfuccacuascsu 2034 AGUAGUGGAUGUCUGCAAAAACC 2747AD-1726083 asgsuggaugUfCfUfgcaaaaaccuL96 1384asdGsgudTudTugcadGaCfauccacusasc 2035 GUAGUGGAUGUCUGCAAAAACCA 2748AD-1726084 gsusggauguCfUfGfcaaaaaccauL96 1385asdTsggdTudTuugcdAgAfcauccacsusa 2036 UAGUGGAUGUCUGCAAAAACCAG 2749AD-1726085 usgsgaugucUfGfCfaaaaaccaguL96 1386asdCsugdGudTuuugdCaGfacauccascsu 2037 AGUGGAUGUCUGCAAAAACCAGA 2750AD-1726086 gsgsaugucuGfCfAfaaaaccagauL96 1387asdTscudGgdTuuuudGcAfgacauccsasc 2038 GUGGAUGUCUGCAAAAACCAGAA 2751AD-1726087 gsasugucugCfAfAfaaaccagaauL96 1388asdTsucdTgdGuuuudTgCfagacaucscsa 2039 UGGAUGUCUGCAAAAACCAGAAG 2752AD-1726090 gsuscugcaaAfAfAfccagaagcguL96 1389asdCsgcdTudCuggudTuUfugcagacsasu 2040 AUGUCUGCAAAAACCAGAAGCGG 2753AD-1726091 uscsugcaaaAfAfCfcagaagcgguL96 1390asdCscgdCudTcuggdTuUfuugcagascsa 2041 UGUCUGCAAAAACCAGAAGCGGC 2754AD-1726092 csusgcaaaaAfCfCfagaagcggcuL96 1391asdGsccdGcdTucugdGuUfuuugcagsasc 2042 GUCUGCAAAAACCAGAAGCGGCA 2755AD-1726095 csasaaaaccAfGfAfagcggcaaauL96 1392asdTsuudGcdCgcuudCuGfguuuuugscsa 2043 UGCAAAAACCAGAAGCGGCAAAA 2756AD-1726096 asasaaaccaGfAfAfgcggcaaaauL96 1393asdTsuudTgdCcgcudTcUfgguuuuusgsc 2044 GCAAAAACCAGAAGCGGCAAAAG 2757AD-1726097 asasaaccagAfAfGfcggcaaaaguL96 1394asdCsuudTudGccgcdTuCfugguuuususg 2045 CAAAAACCAGAAGCGGCAAAAGC 2758AD-1726098 asasaccagaAfGfCfggcaaaagcuL96 1395asdGscudTudTgccgdCuUfcugguuususu 2046 AAAAACCAGAAGCGGCAAAAGCA 2759AD-1726099 asasccagaaGfCfGfgcaaaagcauL96 1396asdTsgcdTudTugccdGcUfucugguususu 2047 AAAACCAGAAGCGGCAAAAGCAG 2760AD-1726103 asgsaagcggCfAfAfaagcagguauL96 1397asdTsacdCudGcuuudTgCfcgcuucusgsg 2048 CCAGAAGCGGCAAAAGCAGGUAC 2761AD-1726113 asasagcaggUfAfCfcugcucacguL96 1398asdCsgudGadGcaggdTaCfcugcuuususg 2049 CAAAAGCAGGUACCUGCUCACGC 2762AD-1726159 csasagugcuGfCfCfcuggcugaauL96 1399asdTsucdAgdCcaggdGcAfgcacuugsasa 2050 UUCAAGUGCUGCCCUGGCUGAAG 2763AD-1726171 usgsgcugaaGfGfAfgaaacuccauL96 1400asdTsggdAgdTuucudCcUfucagccasgsg 2051 CCUGGCUGAAGGAGAAACUCCAA 2764AD-1726184 asascuccaaGfAfUfgaggauuuguL96 1401asdCsaadAudCcucadTcUfuggaguususc 2052 GAAACUCCAAGAUGAGGAUUUGG 2765AD-1726187 uscscaagauGfAfGfgauuuggguuL96 1402asdAsccdCadAauccdTcAfucuuggasgsu 2053 ACUCCAAGAUGAGGAUUUGGGUU 2766AD-1726189 csasagaugaGfGfAfuuuggguuuuL96 1403asdAsaadCcdCaaaudCcUfcaucuugsgsa 2054 UCCAAGAUGAGGAUUUGGGUUUU 2767AD-1726191 asgsaugaggAfUfUfuggguuuucuL96 1404asdGsaadAadCccaadAuCfcucaucususg 2055 CAAGAUGAGGAUUUGGGUUUUCU 2768AD-1726201 gsusgggauuGfAfAfuuaaaacaguL96 1405asdCsugdTudTuaaudTcAfaucccacsgsc 2056 GCGUGGGAUUGAAUUAAAACAGC 2769AD-1726202 usgsggauugAfAfUfuaaaacagcuL96 1406asdGscudGudTuuaadTuCfaaucccascsg 2057 CGUGGGAUUGAAUUAAAACAGCU 2770AD-1726203 gsgsgauugaAfUfUfaaaacagcuuL96 1407asdAsgcdTgdTuuuadAuUfcaaucccsasc 2058 GUGGGAUUGAAUUAAAACAGCUG 2771AD-1726206 asusugaauuAfAfAfacagcugcguL96 1408asdCsgcdAgdCuguudTuAfauucaauscsc 2059 GGAUUGAAUUAAAACAGCUGCGA 2772AD-1726207 ususgaauuaAfAfAfcagcugcgauL96 1409asdTscgdCadGcugudTuUfaauucaasusc 2060 GAUUGAAUUAAAACAGCUGCGAC 2773AD-1726208 usgsaauuaaAfAfCfagcugcgacuL96 1410asdGsucdGcdAgcugdTuUfuaauucasasu 2061 AUUGAAUUAAAACAGCUGCGACA 2774AD-1726209 gsasauuaaaAfCfAfgcugcgacauL96 1411asdTsgudCgdCagcudGuUfuuaauucsasa 2062 UUGAAUUAAAACAGCUGCGACAA 2775AD-1726815 csusggcuUfcUfAfCfccguacccuuL96 1412asAfsgggUfacggguaGfaAfgccagsasa 2063 UUCUGGCUUCUACCCGUACCCUG 2406AD-1726927 cscscuacUfaCfAfAfugugagugauL96 1413asUfscacUfcacauugUfaGfuagggsasg 2064 CUCCCUACUACAAUGUGAGUGAU 2407AD-1726928 cscsuacuAfcAfAfUfgugagugauuL96 1414asAfsucaCfucacauuGfuAfguaggsgsa 2065 UCCCUACUACAAUGUGAGUGAUG 2408AD-1726931 ascsuacaAfuGfUfGfagugaugaguL96 1415asCfsucaUfcacucacAfuUfguagusasg 2066 CUACUACAAUGUGAGUGAUGAGA 2409AD-1726934 ascsaaugUfgAfGfUfgaugagaucuL96 1416asGfsaucUfcaucacuCfaCfauugusasg 2067 CUACAAUGUGAGUGAUGAGAUCU 2410AD-1726935 csasauguGfaGfUfGfaugagaucuuL96 1417asAfsgauCfucaucacUfcAfcauugsusa 2068 UACAAUGUGAGUGAUGAGAUCUC 2411AD-1726936 asasugugAfgUfGfAfugagaucucuL96 1418asGfsagaUfcucaucaCfuCfacauusgsu 2069 ACAAUGUGAGUGAUGAGAUCUCU 2412AD-1726937 asusgugaGfuGfAfUfgagaucucuuL96 1419asAfsgagAfucucaucAfcUfcacaususg 2070 CAAUGUGAGUGAUGAGAUCUCUU 2413AD-1726938 usgsugagUfgAfUfGfagaucucuuuL96 1420asAfsagaGfaucucauCfaCfucacasusu 2071 AAUGUGAGUGAUGAGAUCUCUUU 2414AD-1726939 gsusgaguGfaUfGfAfgaucucuuuuL96 1421asAfsaagAfgaucucaUfcAfcucacsasu 2072 AUGUGAGUGAUGAGAUCUCUUUC 2415AD-1726940 usgsagugAfuGfAfGfaucucuuucuL96 1422asGfsaaaGfagaucucAfuCfacucascsa 2073 UGUGAGUGAUGAGAUCUCUUUCC 2416AD-1726941 gsasgugaUfgAfGfAfucucuuuccuL96 1423asGfsgaaAfgagaucuCfaUfcacucsasc 2074 GUGAGUGAUGAGAUCUCUUUCCA 2417AD-1726942 asgsugauGfaGfAfUfcucuuuccauL96 1424asUfsggaAfagagaucUfcAfucacuscsa 2075 UGAGUGAUGAGAUCUCUUUCCAC 2418AD-1726944 usgsaugaGfaUfCfUfcuuuccacuuL96 1425asAfsgugGfaaagagaUfcUfcaucascsu 2076 AGUGAUGAGAUCUCUUUCCACUG 2420AD-1726952 uscsucuuUfcCfAfCfugcuaugacuL96 1426asGfsucaUfagcagugGfaAfagagasusc 2077 GAUCUCUUUCCACUGCUAUGACG 2423AD-1726961 ascsugcuAfuGfAfCfgguuacacuuL96 1427asAfsgugUfaaccgucAfuAfgcagusgsg 2078 CCACUGCUAUGACGGUUACACUC 2426AD-1727012 csasgacaGfcGfAfUfcugugacaauL96 1428asUfsuguCfacagaucGfc Ufgucugscsc 2079 GGCAGACAGCGAUCUGUGACAAC 2433AD-1727059 csusugaaGfaCfAfGfcgucaccuauL96 1429asUfsaggUfgacgcugUfcUfucaagsgsc 2080 GCCUUGAAGACAGCGUCACCUAC 2441AD-1727140 asasgacuCfcUfUfCfauguacgacuL96 1430asGfsucgUfacaugaaGfgAfgucuusgsg 2081 CCAAGACUCCUUCAUGUACGACA 2451AD-1727142 gsascuccUfuCfAfUfguacgacacuL96 1431asGfsuguCfguacaugAfaGfgagucsusu 2082 AAGACUCCUUCAUGUACGACACC 2452AD-1727181 asgsagacCfaUfAfGfaaggagucguL96 1432asCfsgacUfccuucuaUfgGfucucusgsu 2083 ACAGAGACCAUAGAAGGAGUCGA 2455AD-1727183 asgsaccaUfaGfAfAfggagucgauuL96 1433asAfsucgAfcuccuucUfaUfggucuscsu 2084 AGAGACCAUAGAAGGAGUCGAUG 2457AD-1727184 gsasccauAfgAfAfGfgagucgauguL96 1434asCfsaucGfacuccuuCfuAfuggucsusc 2085 GAGACCAUAGAAGGAGUCGAUGC 2458AD-1727249 usgsaacaUfcUfAfCfcuggugcuauL96 1435asUfsagcAfccagguaGfaUfguucasusg 2086 CAUGAACAUCUACCUGGUGCUAG 2463AD-1727261 usgsgugcUfaGfAfUfggaucagacuL96 1436asGfsucuGfauccaucUfaGfcaccasgsg 2087 CCUGGUGCUAGAUGGAUCAGACA 2472AD-1727263 gsusgcuaGfaUfGfGfaucagacaguL96 1437asCfsuguCfugauccaUfcUfagcacscsa 2088 UGGUGCUAGAUGGAUCAGACAGC 2473AD-1727275 csasacuuCfaCfAfGfgagccaaaauL96 1438asUfsuuuGfgcuccugUfgAfaguugscsu 2089 AGCAACUUCACAGGAGCCAAAAA 2476AD-1727276 asascuucAfcAfGfGfagccaaaaauL96 1439asUfsuuuUfggcuccuGfuGfaaguusgsc 2090 GCAACUUCACAGGAGCCAAAAAG 2477AD-1727278 csusucacAfgGfAfGfccaaaaaguuL96 1440asAfscuuUfuuggcucCfuGfugaagsusu 2091 AACUUCACAGGAGCCAAAAAGUG 2479AD-1727285 gsgsagccAfaAfAfAfgugucuaguuL96 1441asAfscuaGfacacuuuUfuGfgcuccsusg 2092 CAGGAGCCAAAAAGUGUCUAGUC 2486AD-1727286 gsasgccaAfaAfAfGfugucuagucuL96 1442asGfsacuAfgacacuuUfuUfggcucscsu 2093 AGGAGCCAAAAAGUGUCUAGUCA 2487AD-1727288 gscscaaaAfaGfUfGfucuagucaauL96 1443asUfsugaCfuagacacUfuUfuuggcsusc 2094 GAGCCAAAAAGUGUCUAGUCAAC 2489AD-1727289 cscsaaaaAfgUfGfUfcuagucaacuL96 1444asGfsuugAfcuagacaCfuUfuuuggscsu 2095 AGCCAAAAAGUGUCUAGUCAACU 2490AD-1727290 csasaaaaGfuGfUfCfuagucaacuuL96 1445asAfsguuGfacuagacAfcUfuuuugsgsc 2096 GCCAAAAAGUGUCUAGUCAACUU 2491AD-1727291 asasaaagUfgUfCfUfagucaacuuuL96 1446asAfsaguUfgacuagaCfaCfuuuuusgsg 2097 CCAAAAAGUGUCUAGUCAACUUA 2492AD-1727292 asasaaguGfuCfUfAfgucaacuuauL96 1447asUfsaagUfugacuagAfcAfcuuuususg 2098 CAAAAAGUGUCUAGUCAACUUAA 2493AD-1727293 asasagugUfcUfAfGfucaacuuaauL96 1448asUfsuaaGfuugacuaGfaCfacuuususu 2099 AAAAAGUGUCUAGUCAACUUAAU 2494AD-1727298 gsuscuagUfcAfAfCfuuaauugaguL96 1449asCfsucaAfuuaaguuGfaCfuagacsasc 2100 GUGUCUAGUCAACUUAAUUGAGA 2499AD-1727310 usasauugAfgAfAfGfguggcaaguuL96 1450asAfscuuGfccaccuuCfuCfaauuasasg 2101 CUUAAUUGAGAAGGUGGCAAGUU 2501AD-1727318 asasggugGfcAfAfGfuuaugguguuL96 1451asAfscacCfauaacuuGfcCfaccuuscsu 2102 AGAAGGUGGCAAGUUAUGGUGUG 2505AD-1727324 gscsaaguUfaUfGfGfugugaagccuL96 1452asGfsgcuUfcacaccaUfaAfcuugcscsa 2103 UGGCAAGUUAUGGUGUGAAGCCA 2507AD-1727331 asusggugUfgAfAfGfccaagauauuL96 1453asAfsuauCfuuggcuuCfaCfaccausasa 2104 UUAUGGUGUGAAGCCAAGAUAUG 2509AD-1727358 asasaauuUfgGfGfUfcaaagugucuL96 1454asGfsacaCfuuugaccCfaAfauuuusgsg 2105 CCAAAAUUUGGGUCAAAGUGUCU 2511AD-1727359 asasauuuGfgGfUfCfaaagugucuuL96 1455asAfsgacAfcuuugacCfcAfaauuususg 2106 CAAAAUUUGGGUCAAAGUGUCUG 2512AD-1727361 asusuuggGfuCfAfAfagugucugauL96 1456asUfscagAfcacuuugAfcCfcaaaususu 2107 AAAUUUGGGUCAAAGUGUCUGAA 2513AD-1727392 gsusaaugCfaGfAfCfugggucacguL96 1457asCfsgugAfcccagucUfgCfauuacsusg 2108 CAGUAAUGCAGACUGGGUCACGA 2514AD-1727420 asasugaaAfuCfAfAfuuaugaagauL96 1458asUfscuuCfauaauugAfuUfucauusgsa 2109 UCAAUGAAAUCAAUUAUGAAGAC 2518AD-1727427 uscsaauuAfuGfAfAfgaccacaaguL96 1459asCfsuugUfggucuucAfuAfauugasusu 2110 AAUCAAUUAUGAAGACCACAAGU 2522AD-1727428 csasauuaUfgAfAfGfaccacaaguuL96 1460asAfscuuGfuggucuuCfaUfaauugsasu 2111 AUCAAUUAUGAAGACCACAAGUU 2523AD-1727430 asusuaugAfaGfAfCfcacaaguuguL96 1461asCfsaacUfuguggucUfuCfauaaususg 2112 CAAUUAUGAAGACCACAAGUUGA 2525AD-1727431 ususaugaAfgAfCfCfacaaguugauL96 1462asUfscaaCfuugugguCfuUfcauaasusu 2113 AAUUAUGAAGACCACAAGUUGAA 2526AD-1727432 usasugaaGfaCfCfAfcaaguugaauL96 1463asUfsucaAfcuuguggUfcUfucauasasu 2114 AUUAUGAAGACCACAAGUUGAAG 2527AD-1727433 asusgaagAfcCfAfCfaaguugaaguL96 1464asCfsuucAfacuugugGfuCfuucausasa 2115 UUAUGAAGACCACAAGUUGAAGU 2528AD-1727434 usgsaagaCfcAfCfAfaguugaaguuL96 1465asAfscuuCfaacuuguGfgUfcuucasusa 2116 UAUGAAGACCACAAGUUGAAGUC 2529AD-1727435 gsasagacCfaCfAfAfguugaagucuL96 1466asGfsacuUfcaacuugUfgGfucuucsasu 2117 AUGAAGACCACAAGUUGAAGUCA 2530AD-1727436 asasgaccAfcAfAfGfuugaagucauL96 1467asUfsgacUfucaacuuGfuGfgucuuscsa 2118 UGAAGACCACAAGUUGAAGUCAG 2531AD-1727441 csascaagUfuGfAfAfgucagggacuL96 1468asGfsuccCfugacuucAfaCfuugugsgsu 2119 ACCACAAGUUGAAGUCAGGGACU 2535AD-1727442 ascsaaguUfgAfAfGfucagggacuuL96 1469asAfsgucCfcugacuuCfaAfcuugusgsg 2120 CCACAAGUUGAAGUCAGGGACUA 2536AD-1727481 asgsgcagUfgUfAfCfagcaugauguL96 1470asCfsaucAfugcuguaCfaCfugccusgsg 2121 CCAGGCAGUGUACAGCAUGAUGA 2544AD-1727483 gscsagugUfaCfAfGfcaugaugaguL96 1471asCfsucaUfcaugcugUfaCfacugcscsu 2122 AGGCAGUGUACAGCAUGAUGAGC 2545AD-1727565 gsasuggaUfuGfCfAfcaacauggguL96 1472asCfsccaUfguugugcAfaUfccaucsasg 2123 CUGAUGGAUUGCACAACAUGGGC 2548AD-1727566 asusggauUfgCfAfCfaacaugggcuL96 1473asGfscccAfuguugugCfaAfuccauscsa 2124 UGAUGGAUUGCACAACAUGGGCG 2549AD-1727568 gsgsauugCfaCfAfAfcaugggcgguL96 1474asCfscgcCfcauguugUfgCfaauccsasu 2125 AUGGAUUGCACAACAUGGGCGGG 2551AD-1727569 gsascccaAfuUfAfCfugucauugauL96 1475asUfscaaUfgacaguaAfuUfgggucscsc 2126 GGGACCCAAUUACUGUCAUUGAU 2552AD-1727570 ascsccaaUfuAfCfUfgucauugauuL96 1476asAfsucaAfugacaguAfaUfuggguscsc 2127 GGACCCAAUUACUGUCAUUGAUG 2553AD-1727572 cscsaauuAfcUfGfUfcauugaugauL96 1477asUfscauCfaaugacaGfuAfauuggsgsu 2128 ACCCAAUUACUGUCAUUGAUGAG 2554AD-1727584 asusugauGfaGfAfUfccgggacuuuL96 1478asAfsaguCfccggaucUfcAfucaausgsa 2129 UCAUUGAUGAGAUCCGGGACUUG 2558AD-1727612 ususggcaAfgGfAfUfcgcaaaaacuL96 1479asGfsuuuUfugcgaucCfuUfgccaasusg 2130 CAUUGGCAAGGAUCGCAAAAACC 2559AD-1727633 csasagggAfgGfAfUfuaucuggauuL96 1480asAfsuccAfgauaaucCfuCfccuugsgsg 2131 CCCAAGGGAGGAUUAUCUGGAUG 2562AD-1727638 gsasggauUfaUfCfUfggaugucuauL96 1481asUfsagaCfauccagaUfaAfuccucscsc 2132 GGGAGGAUUAUCUGGAUGUCUAU 2563AD-1727639 asgsgauuAfuCfUfGfgaugucuauuL96 1482asAfsuagAfcauccagAfuAfauccuscsc 2133 GGAGGAUUAUCUGGAUGUCUAUG 2564AD-1727640 gsgsauuaUfcUfGfGfaugucuauguL96 1483asCfsauaGfacauccaGfaUfaauccsusc 2134 GAGGAUUAUCUGGAUGUCUAUGU 2565AD-1727641 gsasuuauCfuGfGfAfugucuauguuL96 1484asAfscauAfgacauccAfgAfuaaucscsu 2135 AGGAUUAUCUGGAUGUCUAUGUG 2566AD-1727642 asusuaucUfgGfAfUfgucuauguguL96 1485asCfsacaUfagacaucCfaGfauaauscsc 2136 GGAUUAUCUGGAUGUCUAUGUGU 2567AD-1727643 ususaucuGfgAfUfGfucuauguguuL96 1486asAfscacAfuagacauCfcAfgauaasusc 2137 GAUUAUCUGGAUGUCUAUGUGUU 2568AD-1727644 usasucugGfaUfGfUfcuauguguuuL96 1487asAfsacaCfauagacaUfcCfagauasasu 2138 AUUAUCUGGAUGUCUAUGUGUUU 2569AD-1727645 asuscuggAfuGfUfCfuauguguuuuL96 1488asAfsaacAfcauagacAfuCfcagausasa 2139 UUAUCUGGAUGUCUAUGUGUUUG 2570AD-1727646 uscsuggaUfgUfCfUfauguguuuguL96 1489asCfsaaaCfacauagaCfaUfccagasusa 2140 UAUCUGGAUGUCUAUGUGUUUGG 2571AD-1727663 asasccaaGfuGfAfAfcaucaaugcuL96 1490asGfscauUfgauguucAfcUfugguuscsa 2141 UGAACCAAGUGAACAUCAAUGCU 2573AD-1727664 ascscaagUfgAfAfCfaucaaugcuuL96 1491asAfsgcaUfugauguuCfaCfuuggususc 2142 GAACCAAGUGAACAUCAAUGCUU 2574AD-1727665 cscsaaguGfaAfCfAfucaaugcuuuL96 1492asAfsagcAfuugauguUfcAfcuuggsusu 2143 AACCAAGUGAACAUCAAUGCUUU 2575AD-1727666 csasagugAfaCfAfUfcaaugcuuuuL96 1493asAfsaagCfauugaugUfuCfacuugsgsu 2144 ACCAAGUGAACAUCAAUGCUUUG 2576AD-1727675 asuscaauGfcUfUfUfggcuuccaauL96 1494asUfsuggAfagccaaaGfcAfuugausgsu 2145 ACAUCAAUGCUUUGGCUUCCAAG 2577AD-1727677 csasaugcUfuUfGfGfcuuccaagauL96 1495asUfscuuGfgaagccaAfaGfcauugsasu 2146 AUCAAUGCUUUGGCUUCCAAGAA 2579AD-1727685 usgsgcuuCfcAfAfGfaaagacaauuL96 1496asAfsuugUfcuuucuuGfgAfagccasasa 2147 UUUGGCUUCCAAGAAAGACAAUG 2580AD-1727689 ususccaaGfaAfAfGfacaaugagcuL96 1497asGfscucAfuugucuuUfcUfuggaasgsc 2148 GCUUCCAAGAAAGACAAUGAGCA 2581AD-1727690 uscscaagAfaAfGfAfcaaugagcauL96 1498asUfsgcuCfauugucuUfuCfuuggasasg 2149 CUUCCAAGAAAGACAAUGAGCAA 2582AD-1727693 asasgaaaGfaCfAfAfugagcaacauL96 1499asUfsguuGfcucauugUfcUfuucuusgsg 2150 CCAAGAAAGACAAUGAGCAACAU 2584AD-1727696 asasagacAfaUfGfAfgcaacauguuL96 1500asAfscauGfuugcucaUfuGfucuuuscsu 2151 AGAAAGACAAUGAGCAACAUGUG 2585AD-1727698 asgsacaaUfgAfGfCfaacauguguuL96 1501asAfscacAfuguugcuCfaUfugucususu 2152 AAAGACAAUGAGCAACAUGUGUU 2586AD-1727699 gsascaauGfaGfCfAfacauguguuuL96 1502asAfsacaCfauguugc UfcAfuugucsusu 2153 AAGACAAUGAGCAACAUGUGUUC 2587AD-1727700 ascsaaugAfgCfAfAfcauguguucuL96 1503asGfsaacAfcauguugCfuCfauuguscsu 2154 AGACAAUGAGCAACAUGUGUUCA 2588AD-1727701 csasaugaGfcAfAfCfauguguucauL96 1504asUfsgaaCfacauguuGfc Ufcauugsusc 2155 GACAAUGAGCAACAUGUGUUCAA 2589AD-1727703 asusgagcAfaCfAfUfguguucaaauL96 1505asUfsuugAfacacaugUfuGfcucaususg 2156 CAAUGAGCAACAUGUGUUCAAAG 2590AD-1727705 gsasgcaaCfaUfGfUfguucaaaguuL96 1506asAfscuuUfgaacacaUfgUfugcucsasu 2157 AUGAGCAACAUGUGUUCAAAGUC 2591AD-1727708 csasacauGfuGfUfUfcaaagucaauL96 1507asUfsugaCfuuugaacAfcAfuguugscsu 2158 AGCAACAUGUGUUCAAAGUCAAG 2593AD-1727709 asascaugUfgUfUfCfaaagucaaguL96 1508asCfsuugAfcuuugaaCfaCfauguusgsc 2159 GCAACAUGUGUUCAAAGUCAAGG 2594AD-1727710 ascsauguGfuUfCfAfaagucaagguL96 1509asCfscuuGfacuuugaAfcAfcaugususg 2160 CAACAUGUGUUCAAAGUCAAGGA 2595AD-1727712 asusguguUfcAfAfAfgucaaggauuL96 1510asAfsuccUfugacuuuGfaAfcacausgsu 2161 ACAUGUGUUCAAAGUCAAGGAUA 2596AD-1727713 usgsuguuCfaAfAfGfucaaggauauL96 1511asUfsaucCfuugacuuUfgAfacacasusg 2162 CAUGUGUUCAAAGUCAAGGAUAU 2597AD-1727714 gsusguucAfaAfGfUfcaaggauauuL96 1512asAfsuauCfcuugacuUfuGfaacacsasu 2163 AUGUGUUCAAAGUCAAGGAUAUG 2598AD-1727717 ususcaaaGfuCfAfAfggauauggauL96 1513asUfsccaUfauccuugAfcUfuugaascsa 2164 UGUUCAAAGUCAAGGAUAUGGAA 2599AD-1727718 uscsaaagUfcAfAfGfgauauggaauL96 1514asUfsuccAfuauccuuGfaCfuuugasasc 2165 GUUCAAAGUCAAGGAUAUGGAAA 2600AD-1727821 usasccgaUfuAfCfCfacaagcaacuL96 1515asGfsuugCfuugugguAfaUfcgguascsc 2166 GGUACCGAUUACCACAAGCAACC 2610AD-1727823 cscsgauuAfcCfAfCfaagcaaccauL96 1516asUfsgguUfgcuugugGfuAfaucggsusa 2167 UACCGAUUACCACAAGCAACCAU 2611AD-1727826 asusuaccAfcAfAfGfcaaccaugguL96 1517asCfscauGfguugcuuGfuGfguaauscsg 2168 CGAUUACCACAAGCAACCAUGGC 2613AD-1727829 ascscacaAfgCfAfAfccauggcaguL96 1518asCfsugcCfaugguugCfuUfguggusasa 2169 UUACCACAAGCAACCAUGGCAGG 2616AD-1727883 usgsguguCfuGfAfGfuacuuuguguL96 1519asCfsacaAfaguacucAfgAfcaccascsa 2170 UGUGGUGUCUGAGUACUUUGUGC 2625AD-1727977 gsasagcaGfgAfAfUfuccugaauuuL96 1520asAfsauuCfaggaauuCfcUfgcuucsusu 2171 AAGAAGCAGGAAUUCCUGAAUUU 2629AD-1727978 asasgcagGfaAfUfUfccugaauuuuL96 1521asAfsaauUfcaggaauUfcCfugcuuscsu 2172 AGAAGCAGGAAUUCCUGAAUUUU 2630AD-1727980 gscsaggaAfuUfCfCfugaauuuuauL96 1522asUfsaaaAfuucaggaAfuUfccugcsusu 2173 AAGCAGGAAUUCCUGAAUUUUAU 2631AD-1727981 csasggaaUfuCfCfUfgaauuuuauuL96 1523asAfsuaaAfauucaggAfaUfuccugscsu 2174 AGCAGGAAUUCCUGAAUUUUAUG 2632AD-1727984 gsasauucCfuGfAfAfuuuuaugacuL96 1524asGfsucaUfaaaauucAfgGfaauucscsu 2175 AGGAAUUCCUGAAUUUUAUGACU 2635AD-1727985 asasuuccUfgAfAfUfuuuaugacuuL96 1525asAfsgucAfuaaaauuCfaGfgaauuscsc 2176 GGAAUUCCUGAAUUUUAUGACUA 2636AD-1727986 asusuccuGfaAfUfUfuuaugacuauL96 1526asUfsaguCfauaaaauUfcAfggaaususc 2177 GAAUUCCUGAAUUUUAUGACUAU 2637AD-1727987 ususccugAfaUfUfUfuaugacuauuL96 1527asAfsuagUfcauaaaaUfuCfaggaasusu 2178 AAUUCCUGAAUUUUAUGACUAUG 2638AD-1727989 cscsugaaUfuUfUfAfugacuaugauL96 1528asUfscauAfgucauaaAfaUfucaggsasa 2179 UUCCUGAAUUUUAUGACUAUGAC 2640AD-1727990 csusgaauUfuUfAfUfgacuaugacuL96 1529asGfsucaUfagucauaAfaAfuucagsgsa 2180 UCCUGAAUUUUAUGACUAUGACG 2641AD-1727992 gsasauuuUfaUfGfAfcuaugacguuL96 1530asAfscguCfauagucaUfaAfaauucsasg 2181 CUGAAUUUUAUGACUAUGACGUU 2642AD-1727993 asasuuuuAfuGfAfCfuaugacguuuL96 1531asAfsacgUfcauagucAfuAfaaauuscsa 2182 UGAAUUUUAUGACUAUGACGUUG 2643AD-1727994 asusuuuaUfgAfCfUfaugacguuguL96 1532asCfsaacGfucauaguCfaUfaaaaususc 2183 GAAUUUUAUGACUAUGACGUUGC 2644AD-1727996 ususuaugAfcUfAfUfgacguugccuL96 1533asGfsgcaAfcgucauaGfuCfauaaasasu 2184 AUUUUAUGACUAUGACGUUGCCC 2645AD-1727999 asusgacuAfuGfAfCfguugcccuguL96 1534asCfsaggGfcaacgucAfuAfgucausasa 2185 UUAUGACUAUGACGUUGCCCUGA 2648AD-1728049 csasgacuAfuCfAfGfgcccauuuguL96 1535asCfsaaaUfgggccugAfuAfgucugsgsc 2186 GCCAGACUAUCAGGCCCAUUUGU 2657AD-1728050 asgsacuaUfcAfGfGfcccauuuguuL96 1536asAfscaaAfugggccuGfaUfagucusgsg 2187 CCAGACUAUCAGGCCCAUUUGUC 2658AD-1728061 csgsagggAfaCfAfAfcucgagcuuuL96 1537asAfsagcUfcgaguugUfuCfccucgsgsu 2188 ACCGAGGGAACAACUCGAGCUUU 2662AD-1728062 gsasgggaAfcAfAfCfucgagcuuuuL96 1538asAfsaagCfucgaguuGfuUfcccucsgsg 2189 CCGAGGGAACAACUCGAGCUUUG 2663AD-1728067 asascaacUfcGfAfGfcuuugaggcuL96 1539asGfsccuCfaaagcucGfaGfuuguuscsc 2190 GGAACAACUCGAGCUUUGAGGCU 2666AD-1728085 gscsuuccUfcCfAfAfcuaccacuuuL96 1540asAfsaguGfguaguugGfaGfgaagcscsu 2191 AGGCUUCCUCCAACUACCACUUG 2677AD-1728132 csusgcacAfgGfAfUfaucaaagcuuL96 1541asAfsgcuUfugauaucCfuGfugcagsgsg 2192 CCCUGCACAGGAUAUCAAAGCUC 2685AD-1728137 csasggauAfuCfAfAfagcucuguuuL96 1542asAfsacaGfagcuuugAfuAfuccugsusg 2193 CACAGGAUAUCAAAGCUCUGUUU 2686AD-1728140 gsasuaucAfaAfGfCfucuguuuguuL96 1543asAfscaaAfcagagcuUfuGfauaucscsu 2194 AGGAUAUCAAAGCUCUGUUUGUG 2687AD-1728146 asasagcuCfuGfUfUfugugucugauL96 1544asUfscagAfcacaaacAfgAfgcuuusgsa 2195 UCAAAGCUCUGUUUGUGUCUGAG 2688AD-1728195 asasgaaaGfgCfAfGfcugugagaguL96 1545asCfsucuCfacagcugCfcUfuucuusasu 2196 AUAAGAAAGGCAGCUGUGAGAGA 2696AD-1728204 asgscuguGfaGfAfGfagaugcucauL96 1546asUfsgagCfaucucucUfcAfcagcusgsc 2197 GCAGCUGUGAGAGAGAUGCUCAA 2698AD-1728206 csusgugaGfaGfAfGfaugcucaauuL96 1547asAfsuugAfgcaucucUfcUfcacagscsu 2198 AGCUGUGAGAGAGAUGCUCAAUA 2699AD-1728207 usgsugagAfgAfGfAfugcucaauauL96 1548asUfsauuGfagcaucuCfuCfucacasgsc 2199 GCUGUGAGAGAGAUGCUCAAUAU 2700AD-1728208 gsusgagaGfaGfAfUfgcucaauauuL96 1549asAfsuauUfgagcaucUfcUfcucacsasg 2200 CUGUGAGAGAGAUGCUCAAUAUG 2701AD-1728209 usgsagagAfgAfUfGfcucaauauguL96 1550asCfsauaUfugagcauCfuCfucucascsa 2201 UGUGAGAGAGAUGCUCAAUAUGC 2702AD-1728210 gsasgagaGfaUfGfCfucaauaugcuL96 1551asGfscauAfuugagcaUfcUfcucucsasc 2202 GUGAGAGAGAUGCUCAAUAUGCC 2703AD-1728212 csasggcuAfuGfAfCfaaagucaaguL96 1552asCfsuugAfcuuugucAfuAfgccugsgsg 2203 CCCAGGCUAUGACAAAGUCAAGG 2704AD-1728214 gsgscuauGfaCfAfAfagucaaggauL96 1553asUfsccuUfgacuuugUfcAfuagccsusg 2204 CAGGCUAUGACAAAGUCAAGGAC 2706AD-1728220 gsascaaaGfuCfAfAfggacaucucuL96 1554asGfsagaUfguccuugAfcUfuugucsasu 22056 AUGACAAAGUCAAGGACAUCUCA 2707AD-1728244 ususguacUfgGfAfGfgagugagucuL96 1555asGfsacuCfacuccucCfaGfuacaasasg 2206 CUUUGUACUGGAGGAGUGAGUCC 2713AD-1728258 asasuacuUfgCfAfGfaggugauucuL96 1556asGfsaauCfaccucugCfaAfguauusgsg 2207 CCAAUACUUGCAGAGGUGAUUCU 2722AD-1728260 usascuugCfaGfAfGfgugauucuguL96 1557asCfsagaAfucaccucUfgCfaaguasusu 2208 AAUACUUGCAGAGGUGAUUCUGG 2723AD-1728269 usgsauagUfuCfAfCfaagagaaguuL96 1558asAfscuuCfucuugugAfaCfuaucasasg 2209 CUUGAUAGUUCACAAGAGAAGUC 2726AD-1728270 gsasuaguUfcAfCfAfagagaagucuL96 1559asGfsacuUfcucuuguGfaAfcuaucsasa 2210 UUGAUAGUUCACAAGAGAAGUCG 2727AD-1728271 asusaguuCfaCfAfAfgagaagucguL96 1560asCfsgacUfucucuugUfgAfacuauscsa 2211 UGAUAGUUCACAAGAGAAGUCGU 2728AD-1728272 usasguucAfcAfAfGfagaagucguuL96 1561asAfscgaCfuucucuuGfuGfaacuasusc 2212 GAUAGUUCACAAGAGAAGUCGUU 2729AD-1728273 asgsuucaCfaAfGfAfgaagucguuuL96 1562asAfsacgAfcuucucuUfgUfgaacusasu 2213 AUAGUUCACAAGAGAAGUCGUUU 2730AD-1728274 gsusucacAfaGfAfGfaagucguuuuL96 1563asAfsaacGfacuucucUfuGfugaacsusa 2214 UAGUUCACAAGAGAAGUCGUUUC 2731AD-1728275 ususcacaAfgAfGfAfagucguuucuL96 1564asGfsaaaCfgacuucuCfuUfgugaascsu 2215 AGUUCACAAGAGAAGUCGUUUCA 2732AD-1728276 uscsacaaGfaGfAfAfgucguuucauL96 1565asUfsgaaAfcgacuucUfcUfugugasasc 2216 GUUCACAAGAGAAGUCGUUUCAU 2733AD-1728277 csascaagAfgAfAfGfucguuucauuL96 1566asAfsugaAfacgacuuCfuCfuugugsasa 2217 UUCACAAGAGAAGUCGUUUCAUU 2734AD-1728278 ascsaagaGfaAfGfUfcguuucauuuL96 1567asAfsaugAfaacgacuUfcUfcuugusgsa 2218 UCACAAGAGAAGUCGUUUCAUUC 2735AD-1728279 csasagagAfaGfUfCfguuucauucuL96 1568asGfsaauGfaaacgacUfuCfucuugsusg 2219 CACAAGAGAAGUCGUUUCAUUCA 2736AD-1728280 asasgagaAfgUfCfGfuuucauucauL96 1569asUfsgaaUfgaaacgaCfuUfcucuusgsu 2220 ACAAGAGAAGUCGUUUCAUUCAA 2737AD-1728282 gsasgaagUfcGfUfUfucauucaaguL96 1570asCfsuugAfaugaaacGfaCfuucucsusu 2221 AAGAGAAGUCGUUUCAUUCAAGU 2739AD-1728283 asgsaaguCfgUfUfUfcauucaaguuL96 1571asAfscuuGfaaugaaaCfgAfcuucuscsu 2222 AGAGAAGUCGUUUCAUUCAAGUU 2740AD-1728284 gsasagucGfuUfUfCfauucaaguuuL96 1572asAfsacuUfgaaugaaAfcGfacuucsusc 2223 GAGAAGUCGUUUCAUUCAAGUUG 2741AD-1728285 asasgucgUfuUfCfAfuucaaguuguL96 1573asCfsaacUfugaaugaAfaCfgacuuscsu 2224 AGAAGUCGUUUCAUUCAAGUUGG 2742AD-1728286 asgsucguUfuCfAfUfucaaguugguL96 1574asCfscaaCfuugaaugAfaAfcgacususc 2225 GAAGUCGUUUCAUUCAAGUUGGU 2743AD-1728300 gsasguagUfgGfAfUfgucugcaaauL96 1575asUfsuugCfagacaucCfaCfuacucscsc 2226 GGGAGUAGUGGAUGUCUGCAAAA 2744AD-1728301 asgsuaguGfgAfUfGfucugcaaaauL96 1576asUfsuuuGfcagacauCfcAfcuacuscsc 2227 GGAGUAGUGGAUGUCUGCAAAAA 2745AD-1728302 gsusagugGfaUfGfUfcugcaaaaauL96 1577asUfsuuuUfgcagacaUfcCfacuacsusc 2228 GAGUAGUGGAUGUCUGCAAAAAC 2746AD-1728303 usasguggAfuGfUfCfugcaaaaacuL96 1578asGfsuuuUfugcagacAfuCfcacuascsu 2229 AGUAGUGGAUGUCUGCAAAAACC 2747AD-1728307 gsgsauguCfuGfCfAfaaaaccagauL96 1579asUfscugGfuuuuugcAfgAfcauccsasc 2230 GUGGAUGUCUGCAAAAACCAGAA 2751AD-1728308 gsasugucUfgCfAfAfaaaccagaauL96 1580asUfsucuGfguuuuugCfaGfacaucscsa 2231 UGGAUGUCUGCAAAAACCAGAAG 2752AD-1728311 gsuscugcAfaAfAfAfccagaagcguL96 1581asCfsgcuUfcugguuuUfuGfcagacsasu 2232 AUGUCUGCAAAAACCAGAAGCGG 2753AD-1728312 uscsugcaAfaAfAfCfcagaagcgguL96 1582asCfscgcUfucugguuUfuUfgcagascsa 2233 UGUCUGCAAAAACCAGAAGCGGC 2754AD-1728317 asasaaacCfaGfAfAfgcggcaaaauL96 1583asUfsuuuGfccgcuucUfgGfuuuuusgsc 2234 GCAAAAACCAGAAGCGGCAAAAG 2757AD-1728318 asasaaccAfgAfAfGfcggcaaaaguL96 1584asCfsuuuUfgccgcuuCfuGfguuuususg 2235 CAAAAACCAGAAGCGGCAAAAGC 2758AD-1728320 asasccagAfaGfCfGfgcaaaagcauL96 1585asUfsgcuUfuugccgcUfuCfugguususu 2236 AAAACCAGAAGCGGCAAAAGCAG 2760AD-1728324 asgsaagcGfgCfAfAfaagcagguauL96 1586asUfsaccUfgcuuuugCfcGfcuucusgsg 2237 CCAGAAGCGGCAAAAGCAGGUAC 2761AD-1728405 asascuccAfaGfAfUfgaggauuuguL96 1587asCfsaaaUfccucaucUfuGfgaguususc 2238 GAAACUCCAAGAUGAGGAUUUGG 2765AD-1728408 uscscaagAfuGfAfGfgauuuggguuL96 1588asAfscccAfaauccucAfuCfuuggasgsu 2239 ACUCCAAGAUGAGGAUUUGGGUU 2766AD-1728410 csasagauGfaGfGfAfuuuggguuuuL96 1589asAfsaacCfcaaauccUfcAfucuugsgsa 2240 UCCAAGAUGAGGAUUUGGGUUUU 2767AD-1728412 asgsaugaGfgAfUfUfuggguuuucuL96 1590asGfsaaaAfcccaaauCfcUfcaucususg 2241 CAAGAUGAGGAUUUGGGUUUUCU 2768AD-1728422 gsusgggaUfuGfAfAfuuaaaacaguL96 1591asCfsuguUfuuaauucAfaUfcccacsgsc 2242 GCGUGGGAUUGAAUUAAAACAGC 2769AD-1728423 usgsggauUfgAfAfUfuaaaacagcuL96 1592asGfscugUfuuuaauuCfaAfucccascsg 2243 CGUGGGAUUGAAUUAAAACAGCU 2770AD-1728424 gsgsgauuGfaAfUfUfaaaacagcuuL96 1593asAfsgcuGfuuuuaauUfcAfaucccsasc 2244 GUGGGAUUGAAUUAAAACAGCUG 2771AD-1728427 asusugaaUfuAfAfAfacagcugcguL96 1594asCfsgcaGfcuguuuuAfaUfucaauscsc 2245 GGAUUGAAUUAAAACAGCUGCGA 2772AD-1728447 asasgggaAfuGfUfGfaccaggucuuL96 1595asAfsgadCc(Tgn)ggucacAfuUfcccuuscsc 2246 GGAAGGGAAUGUGACCAGGUCUA 2385AD-1728461 asgsgucuAfgGfUfCfuggaguuucuL96 1596asGfsaadAc(Tgn)ccagacCfuAfgaccusgsg 2247 CCAGGUCUAGGUCUGGAGUUUCA 2391AD-1728470 uscsuggaGfuUfUfCfagcuuggacuL96 1597asGfsucdCa(Agn)gcugaaAfcUfccagascsc 2248 GGUCUGGAGUUUCAGCUUGGACA 2393AD-1728471 csusggagUfuUfCfAfgcuuggacauL96 1598asUfsgudCc(Agn)agcugaAfaCfuccagsasc 2249 GUCUGGAGUUUCAGCUUGGACAC 2394AD-1728659 uscscuucUfgGfCfUfucuacccguuL96 1599asAfscgdGg(Tgn)agaagcCfaGfaaggascsa 2250 UGUCCUUCUGGCUUCUACCCGUA 2397AD-1728664 csusggcuUfcUfAfCfccguacccuuL96 1412asAfsggdGu(Agn)cggguaGfaAfgccagsasa 2251 UUCUGGCUUCUACCCGUACCCUG 2400AD-1728671 csusacccGfuAfCfCfcugugcagauL96 1600asUfscudGc(Agn)caggguAfcGfgguagsasa 2252 UUCUACCCGUACCCUGUGCAGAC 2401AD-1728685 usgscagaCfaCfGfUfaccugcagauL96 1601asUfscudGc(Agn)gguacgUfgUfcugcascsa 2253 UGUGCAGACACGUACCUGCAGAU 2402AD-1728736 asasggcaGfaGfUfGfcagagcaauuL96 1602asAfsuudGc(Tgn)cugcacUfcUfgccuuscsc 2254 GGAAGGCAGAGUGCAGAGCAAUC 2403AD-1728777 cscsuacuAfcAfAfUfgugagugauuL96 1414asAfsucdAc(Tgn)cacauuGfuAfguaggsgsa 2255 UCCCUACUACAAUGUGAGUGAUG 2407AD-1728784 csasauguGfaGfUfGfaugagaucuuL96 1417asAfsgadTc(Tgn)caucacUfcAfcauugsusa 2256 UACAAUGUGAGUGAUGAGAUCUC 2411AD-1728786 asusgugaGfuGfAfUfgagaucucuuL96 1419asAfsgadGa(Tgn)cucaucAfcUfcacaususg 2257 CAAUGUGAGUGAUGAGAUCUCUU 2413AD-1728787 usgsugagUfgAfUfGfagaucucuuuL96 1420asAfsagdAg(Agn)ucucauCfaCfucacasusu 2258 AAUGUGAGUGAUGAGAUCUCUUU 2414AD-1728789 usgsagugAfuGfAfGfaucucuuucuL96 1422asGfsaadAg(Agn)gaucucAfuCfacucascsa 2259 UGUGAGUGAUGAGAUCUCUUUCC 2416AD-1728793 usgsaugaGfaUfCfUfcuuuccacuuL96 1425asAfsgudGg(Agn)aagagaUfcUfcaucascsu 2260 AGUGAUGAGAUCUCUUUCCACUG 2420AD-1728801 uscsucuuUfcCfAfCfugcuaugacuL96 1426asGfsucdAu(Agn)gcagugGfaAfagagasusc 2261 GAUCUCUUUCCACUGCUAUGACG 2423AD-1728802 csuscuuuCfcAfCfUfgcuaugacguL96 1603asCfsgudCa(Tgn)agcaguGfgAfaagagsasu 2262 AUCUCUUUCCACUGCUAUGACGG 2424AD-1728810 ascsugcuAfuGfAfCfgguuacacuuL96 1427asAfsgudGu(Agn)accgucAfuAfgcagusgsg 2263 CCACUGCUAUGACGGUUACACUC 2426AD-1728811 csusgcuaUfgAfCfGfguuacacucuL96 1604asGfsagdTg(Tgn)aaccguCfaUfagcagsusg 2264 CACUGCUAUGACGGUUACACUCU 2427AD-1728827 uscsgcacCfuGfCfCfaagugaauguL96 1605asCfsaudTc(Agn)cuuggcAfgGfugcgasusu 2265 AAUCGCACCUGCCAAGUGAAUGG 2431AD-1728861 csasgacaGfcGfAfUfcugugacaauL96 1428asUfsugdTc(Agn)cagaucGfcUfgucugscsc 2266 GGCAGACAGCGAUCUGUGACAAC 2433AD-1728863 gsascagcGfaUfCfUfgugacaacguL96 1606asCfsgudTg(Tgn)cacagaUfcGfcugucsusg 2267 CAGACAGCGAUCUGUGACAACGG 2435AD-1728877 usgsgcacAfaGfGfAfaggugggcauL96 1607asUfsgcdCc(Agn)ccuuccUfuGfugccasasu 2268 AUUGGCACAAGGAAGGUGGGCAG 2439AD-1728909 ususgaagAfcAfGfCfgucaccuacuL96 1608asGfsuadGg(Tgn)gacgcuGfuCfuucaasgsg 2269 CCUUGAAGACAGCGUCACCUACC 2442AD-1728990 asgsacucCfuUfCfAfuguacgacauL96 1609asUfsgudCg(Tgn)acaugaAfgGfagucususg 2270 CAAGACUCCUUCAUGUACGACAC 2451AD-1728995 csasagagGfuGfGfCfcgaagcuuuuL96 1610asAfsaadGc(Tgn)ucggccAfcCfucuugsasg 2271 CUCAAGAGGUGGCCGAAGCUUUC 2453AD-1729031 gsasgaccAfuAfGfAfaggagucgauL96 1611asUfscgdAc(Tgn)ccuucuAfuGfgucucsusg 2272 CAGAGACCAUAGAAGGAGUCGAU 2456AD-1729089 csasggcuCfcAfUfGfaacaucuacuL96 1612asGfsuadGa(Tgn)guucauGfgAfgccugsasa 2273 UUCAGGCUCCAUGAACAUCUACC 2462AD-1729103 asuscuacCfuGfGfUfgcuagaugguL96 1613asCfscadTc(Tgn)agcaccAfgGfuagausgsu 2274 ACAUCUACCUGGUGCUAGAUGGA 2466AD-1729105 csusaccuGfgUfGfCfuagauggauuL96 1614asAfsucdCa(Tgn)cuagcaCfcAfgguagsasu 2275 AUCUACCUGGUGCUAGAUGGAUC 2468AD-1729106 usasccugGfuGfCfUfagauggaucuL96 1615asGfsaudCc(Agn)ucuagcAfcCfagguasgsa 2276 UCUACCUGGUGCUAGAUGGAUCA 2469AD-1729110 usgsgugcUfaGfAfUfggaucagacuL96 1436asGfsucdTg(Agn)uccaucUfaGfcaccasgsg 2277 CCUGGUGCUAGAUGGAUCAGACA 2472AD-1729112 gsusgcuaGfaUfGfGfaucagacaguL96 1437asCfsugdTc(Tgn)gauccaUfcUfagcacscsa 2278 UGGUGCUAGAUGGAUCAGACAGC 2473AD-1729130 csascaggAfgCfCfAfaaaagugucuL96 1616asGfsacdAc(Tgn)uuuuggCfuCfcugugsasa 2279 UUCACAGGAGCCAAAAAGUGUCU 2482AD-1729132 csasggagCfcAfAfAfaagugucuauL96 1617asUfsagdAc(Agn)cuuuuuGfgCfuccugsusg 2280 CACAGGAGCCAAAAAGUGUCUAG 2484AD-1729134 gsgsagccAfaAfAfAfgugucuaguuL96 1441asAfscudAg(Agn)cacuuuUfuGfgcuccsusg 2281 CAGGAGCCAAAAAGUGUCUAGUC 2486AD-1729136 asgsccaaAfaAfGfUfgucuagucauL96 1618asUfsgadCu(Agn)gacacuUfuUfuggcuscsc 2282 GGAGCCAAAAAGUGUCUAGUCAA 2488AD-1729137 gscscaaaAfaGfUfGfucuagucaauL96 1443asUfsugdAc(Tgn)agacacUfuUfuuggcsusc 2283 GAGCCAAAAAGUGUCUAGUCAAC 2489AD-1729139 csasaaaaGfuGfUfCfuagucaacuuL96 1445asAfsgudTg(Agn)cuagacAfcUfuuuugsgsc 2284 GCCAAAAAGUGUCUAGUCAACUU 2491AD-1729141 asasaaguGfuCfUfAfgucaacuuauL96 1447asUfsaadGu(Tgn)gacuagAfcAfcuuuususg 2285 CAAAAAGUGUCUAGUCAACUUAA 2493AD-1729142 asasagugUfcUfAfGfucaacuuaauL96 1448asUfsuadAg(Tgn)ugacuaGfaCfacuuususu 2286 AAAAAGUGUCUAGUCAACUUAAU 2494AD-1729151 asgsucaaCfuUfAfAfuugagaagguL96 1619asCfscudTc(Tgn)caauuaAfgUfugacusasg 2287 CUAGUCAACUUAAUUGAGAAGGU 2500AD-1729180 asusggugUfgAfAfGfccaagauauuL96 1453asAfsuadTc(Tgn)uggcuuCfaCfaccausasa 2288 UUAUGGUGUGAAGCCAAGAUAUG 2509AD-1729207 asasaauuUfgGfGfUfcaaagugucuL96 1454asGfsacdAc(Tgn)uugaccCfaAfauuuusgsg 2289 CCAAAAUUUGGGUCAAAGUGUCU 2511AD-1729242 usasaugcAfgAfCfUfgggucacgauL96 1620asUfscgdTg(Agn)cccaguCfuGfcauuascsu 2290 AGUAAUGCAGACUGGGUCACGAA 2515AD-1729269 asasugaaAfuCfAfAfuuaugaagauL96 1458asUfscudTc(Agn)uaauugAfuUfucauusgsa 2291 UCAAUGAAAUCAAUUAUGAAGAC 2518AD-1729271 usgsaaauCfaAfUfUfaugaagaccuL96 1621asGfsgudCu(Tgn)cauaauUfgAfuuucasusu 2292 AAUGAAAUCAAUUAUGAAGACCA 2519AD-1729274 asasucaaUfuAfUfGfaagaccacauL96 1622asUfsgudGg(Tgn)cuucauAfaUfugauususc 2293 GAAAUCAAUUAUGAAGACCACAA 2520AD-1729277 csasauuaUfgAfAfGfaccacaaguuL96 1460asAfscudTg(Tgn)ggucuuCfaUfaauugsasu 2294 AUCAAUUAUGAAGACCACAAGUU 2523AD-1729280 ususaugaAfgAfCfCfacaaguugauL96 1462asUfscadAc(Tgn)ugugguCfuUfcauaasusu 2295 AAUUAUGAAGACCACAAGUUGAA 2526AD-1729285 asasgaccAfcAfAfGfuugaagucauL96 1467asUfsgadCu(Tgn)caacuuGfuGfgucuuscsa 2296 UGAAGACCACAAGUUGAAGUCAG 2531AD-1729288 ascscacaAfgUfUfGfaagucaggguL96 1623asCfsccdTg(Agn)cuucaaCfuUfgugguscsu 2297 AGACCACAAGUUGAAGUCAGGGA 2533AD-1729290 csascaagUfuGfAfAfgucagggacuL96 1468asGfsucdCc(Tgn)gacuucAfaCfuugugsgsu 2298 ACCACAAGUUGAAGUCAGGGACU 2535AD-1729296 ususgaagUfcAfGfGfgacuaacacuL96 1624asGfsugdTu(Agn)gucccuGfaCfuucaascsu 2299 AGUUGAAGUCAGGGACUAACACC 2538AD-1729297 usgsaaguCfaGfGfGfacuaacaccuL96 1625asGfsgudGu(Tgn)agucccUfgAfcuucasasc 2300 GUUGAAGUCAGGGACUAACACCA 2539AD-1729300 asgsucagGfgAfCfUfaacaccaaguL96 1626asCfsuudGg(Tgn)guuaguCfcCfugacususc 2301 GAAGUCAGGGACUAACACCAAGA 2540AD-1729413 usgsauggAfuUfGfCfacaacaugguL96 1627asCfscadTg(Tgn)ugugcaAfuCfcaucasgsu 2302 ACUGAUGGAUUGCACAACAUGGG 2547AD-1729461 ususggcaAfgGfAfUfcgcaaaaacuL96 1479asGfsuudTu(Tgn)gcgaucCfuUfgccaasusg 2303 CAUUGGCAAGGAUCGCAAAAACC 2559AD-1729462 usgsgcaaGfgAfUfCfgcaaaaaccuL96 1628asGfsgudTu(Tgn)ugcgauCfcUfugccasasu 2304 AUUGGCAAGGAUCGCAAAAACCC 2560AD-1729463 gsgscaagGfaUfCfGfcaaaaacccuL96 1629asGfsggdTu(Tgn)uugcgaUfcCfuugccsasa 2305 UUGGCAAGGAUCGCAAAAACCCA 2561AD-1729487 gsasggauUfaUfCfUfggaugucuauL96 1481asUfsagdAc(Agn)uccagaUfaAfuccucscsc 2306 GGGAGGAUUAUCUGGAUGUCUAU 2563AD-1729514 cscsaaguGfaAfCfAfucaaugcuuuL96 1492asAfsagdCa(Tgn)ugauguUfcAfcuuggsusu 2307 AACCAAGUGAACAUCAAUGCUUU 2575AD-1729515 csasagugAfaCfAfUfcaaugcuuuuL96 1493asAfsaadGc(Agn)uugaugUfuCfacuugsgsu 2308 ACCAAGUGAACAUCAAUGCUUUG 2576AD-1729524 asuscaauGfcUfUfUfggcuuccaauL96 1494asUfsugdGa(Agn)gccaaaGfcAfuugausgsu 2309 ACAUCAAUGCUUUGGCUUCCAAG 2577AD-1729525 uscsaaugCfuUfUfGfgcuuccaaguL96 1630asCfsuudGg(Agn)agccaaAfgCfauugasusg 2310 CAUCAAUGCUUUGGCUUCCAAGA 2578AD-1729538 ususccaaGfaAfAfGfacaaugagcuL96 1497asGfscudCa(Tgn)ugucuuUfcUfuggaasgsc 2311 GCUUCCAAGAAAGACAAUGAGCA 2581AD-1729539 uscscaagAfaAfGfAfcaaugagcauL96 1498asUfsgcdTc(Agn)uugucuUfuCfuuggasasg 2312 CUUCCAAGAAAGACAAUGAGCAA 2582AD-1729541 csasagaaAfgAfCfAfaugagcaacuL96 1631asGfsuudGc(Tgn)cauuguCfuUfucuugsgsa 2313 UCCAAGAAAGACAAUGAGCAACA 2583AD-1729545 asasagacAfaUfGfAfgcaacauguuL96 1500asAfscadTg(Tgn)ugcucaUfuGfucuuuscsu 2314 AGAAAGACAAUGAGCAACAUGUG 2585AD-1729548 gsascaauGfaGfCfAfacauguguuuL96 1502asAfsacdAc(Agn)uguugcUfcAfuugucsusu 2315 AAGACAAUGAGCAACAUGUGUUC 2587AD-1729550 csasaugaGfcAfAfCfauguguucauL96 1504asUfsgadAc(Agn)cauguuGfcUfcauugsusc 2316 GACAAUGAGCAACAUGUGUUCAA 2589AD-1729552 asusgagcAfaCfAfUfguguucaaauL96 1505asUfsuudGa(Agn)cacaugUfuGfcucaususg 2317 CAAUGAGCAACAUGUGUUCAAAG 2590AD-1729555 asgscaacAfuGfUfGfuucaaagucuL96 1632asGfsacdTu(Tgn)gaacacAfuGfuugcuscsa 2318 UGAGCAACAUGUGUUCAAAGUCA 2592AD-1729557 csasacauGfuGfUfUfcaaagucaauL96 1507asUfsugdAc(Tgn)uugaacAfcAfuguugscsu 2319 AGCAACAUGUGUUCAAAGUCAAG 2593AD-1729559 ascsauguGfuUfCfAfaagucaagguL96 1509asCfscudTg(Agn)cuuugaAfcAfcaugususg 2320 CAACAUGUGUUCAAAGUCAAGGA 2595AD-1729561 asusguguUfcAfAfAfgucaaggauuL96 1510asAfsucdCu(Tgn)gacuuuGfaAfcacausgsu 2321 ACAUGUGUUCAAAGUCAAGGAUA 2596AD-1729562 usgsuguuCfaAfAfGfucaaggauauL96 1511asUfsaudCc(Tgn)ugacuuUfgAfacacasusg 2322 CAUGUGUUCAAAGUCAAGGAUAU 2597AD-1729567 uscsaaagUfcAfAfGfgauauggaauL96 1514asUfsucdCa(Tgn)auccuuGfaCfuuugasasc 2323 GUUCAAAGUCAAGGAUAUGGAAA 2600AD-1729568 csasaaguCfaAfGfGfauauggaaauL96 1633asUfsuudCc(Agn)uauccuUfgAfcuuugsasa 2324 UUCAAAGUCAAGGAUAUGGAAAA 2601AD-1729619 usgsaaagCfcAfGfUfcucugagucuL96 1634asGfsacdTc(Agn)gagacuGfgCfuuucasusc 2325 GAUGAAAGCCAGUCUCUGAGUCU 2602AD-1729643 usgsgcauGfgUfUfUfgggaacacauL96 1635asUfsgudGu(Tgn)cccaaaCfcAfugccascsa 2326 UGUGGCAUGGUUUGGGAACACAG 2607AD-1729667 gsgsguacCfgAfUfUfaccacaagcuL96 1636asGfscudTg(Tgn)gguaauCfgGfuacccsusu 2327 AAGGGUACCGAUUACCACAAGCA 2609AD-1729670 usasccgaUfuAfCfCfacaagcaacuL96 1515asGfsuudGc(Tgn)ugugguAfaUfcgguascsc 2328 GGUACCGAUUACCACAAGCAACC 2610AD-1729673 csgsauuaCfcAfCfAfagcaaccauuL96 1637asAfsugdGu(Tgn)gcuuguGfgUfaaucgsgsu 2329 ACCGAUUACCACAAGCAACCAUG 2612AD-1729677 usasccacAfaGfCfAfaccauggcauL96 1638asUfsgcdCa(Tgn)gguugcUfuGfugguasasu 2330 AUUACCACAAGCAACCAUGGCAG 2615AD-1729688 ascscaugGfcAfGfGfccaagaucuuL96 1639asAfsgadTc(Tgn)uggccuGfcCfauggususg 2331 CAACCAUGGCAGGCCAAGAUCUC 2618AD-1729690 csasuggcAfgGfCfCfaagaucucauL96 1640asUfsgadGa(Tgn)cuuggcCfuGfccaugsgsu 2332 ACCAUGGCAGGCCAAGAUCUCAG 2620AD-1729729 csusguggUfgUfCfUfgaguacuuuuL96 1641asAfsaadGu(Agn)cucagaCfaCfcacagscsc 2333 GGCUGUGGUGUCUGAGUACUUUG 2622AD-1729802 asgscgggAfcCfUfGfgagauagaauL96 1642asUfsucdTa(Tgn)cuccagGfuCfccgcususc 2334 GAAGCGGGACCUGGAGAUAGAAG 2628AD-1729841 gsasagcaGfgAfAfUfuccugaauuuL96 1520asAfsaudTc(Agn)ggaauuCfcUfgcuucsusu 2335 AAGAAGCAGGAAUUCCUGAAUUU 2629AD-1729849 asasuuccUfgAfAfUfuuuaugacuuL96 1525asAfsgudCa(Tgn)aaaauuCfaGfgaauuscsc 2336 GGAAUUCCUGAAUUUUAUGACUA 2636AD-1729850 asusuccuGfaAfUfUfuuaugacuauL96 1526asUfsagdTc(Agn)uaaaauUfcAfggaaususc 2337 GAAUUCCUGAAUUUUAUGACUAU 2637AD-1729852 uscscugaAfuUfUfUfaugacuauguL96 1643asCfsaudAg(Tgn)cauaaaAfuUfcaggasasu 2338 AUUCCUGAAUUUUAUGACUAUGA 2639AD-1729854 csusgaauUfuUfAfUfgacuaugacuL96 1529asGfsucdAu(Agn)gucauaAfaAfuucagsgsa 2339 UCCUGAAUUUUAUGACUAUGACG 2641AD-1729856 gsasauuuUfaUfGfAfcuaugacguuL96 1530asAfscgdTc(Agn)uagucaUfaAfaauucsasg 2340 CUGAAUUUUAUGACUAUGACGUU 2642AD-1729861 ususaugaCfuAfUfGfacguugcccuL96 1644asGfsggdCa(Agn)cgucauAfgUfcauaasasa 2341 UUUUAUGACUAUGACGUUGCCCU 2646AD-1729862 usasugacUfaUfGfAfcguugcccuuL96 1645asAfsggdGc(Agn)acgucaUfaGfucauasasa 2342 UUUAUGACUAUGACGUUGCCCUG 2647AD-1729869 asusgacgUfuGfCfCfcugaucaaguL96 1646asCfsuudGa(Tgn)cagggcAfaCfgucausasg 2343 CUAUGACGUUGCCCUGAUCAAGC 2651AD-1729870 usgsacguUfgCfCfCfugaucaagcuL96 1647asGfscudTg(Agn)ucagggCfaAfcgucasusa 2344 UAUGACGUUGCCCUGAUCAAGCU 2652AD-1729872 ascsguugCfcCfUfGfaucaagcucuL96 1648asGfsagdCu(Tgn)gaucagGfgCfaacguscsa 2345 UGACGUUGCCCUGAUCAAGCUCA 2654AD-1729926 gsasgggaAfcAfAfCfucgagcuuuuL96 1538asAfsaadGc(Tgn)cgaguuGfuUfcccucsgsg 2346 CCGAGGGAACAACUCGAGCUUUG 2663AD-1729933 csasacucGfaGfCfUfuugaggcuuuL96 1649asAfsagdCc(Tgn)caaagcUfcGfaguugsusu 2347 AACAACUCGAGCUUUGAGGCUUC 2667AD-1729941 gscsuuugAfgGfCfUfuccuccaacuL96 1650asGfsuudGg(Agn)ggaagcCfuCfaaagcsusc 2348 GAGCUUUGAGGCUUCCUCCAACU 2672AD-1729947 asgsgcuuCfcUfCfCfaacuaccacuL96 1651asGfsugdGu(Agn)guuggaGfgAfagccuscsa 2349 UGAGGCUUCCUCCAACUACCACU 2676AD-1729951 ususccucCfaAfCfUfaccacuugcuL96 1652asGfscadAg(Tgn)gguaguUfgGfaggaasgsc 2350 GCUUCCUCCAACUACCACUUGCC 2678AD-1729992 csuscccuGfcAfCfAfggauaucaauL96 1653asUfsugdAu(Agn)uccuguGfcAfgggagscsa 2351 UGCUCCCUGCACAGGAUAUCAAA 2682AD-1729993 uscsccugCfaCfAfGfgauaucaaauL96 1654asUfsuudGa(Tgn)auccugUfgCfagggasgsc 2352 GCUCCCUGCACAGGAUAUCAAAG 2683AD-1729994 cscscugcAfcAfGfGfauaucaaaguL96 1655asCfsuudTg(Agn)uauccuGfuGfcagggsasg 2353 CUCCCUGCACAGGAUAUCAAAGC 2684AD-1729996 csusgcacAfgGfAfUfaucaaagcuuL96 1541asAfsgcdTu(Tgn)gauaucCfuGfugcagsgsg 2354 CCCUGCACAGGAUAUCAAAGCUC 2685AD-1730001 csasggauAfuCfAfAfagcucuguuuL96 1542asAfsacdAg(Agn)gcuuugAfuAfuccugsusg 2355 CACAGGAUAUCAAAGCUCUGUUU 2686AD-1730042 gscsugacUfcGfGfAfaggaggucuuL96 1656asAfsgadCc(Tgn)ccuuccGfaGfucagcsusu 2356 AAGCUGACUCGGAAGGAGGUCUA 2689AD-1730048 uscsggaaGfgAfGfGfucuacaucauL96 1657asUfsgadTg(Tgn)agaccuCfcUfuccgasgsu 2357 ACUCGGAAGGAGGUCUACAUCAA 2692AD-1730053 asgsgaggUfcUfAfCfaucaagaauuL96 1658asAfsuudCu(Tgn)gauguaGfaCfcuccususc 2358 GAAGGAGGUCUACAUCAAGAAUG 2695AD-1730059 asasgaaaGfgCfAfGfcugugagaguL96 1545asCfsucdTc(Agn)cagcugCfcUfuucuusasu 2359 AUAAGAAAGGCAGCUGUGAGAGA 2696AD-1730068 asgscuguGfaGfAfGfagaugcucauL96 1546asUfsgadGc(Agn)ucucucUfcAfcagcusgsc 2360 GCAGCUGUGAGAGAGAUGCUCAA 2698AD-1730071 usgsugagAfgAfGfAfugcucaauauL96 1548asUfsaudTg(Agn)gcaucuCfuCfucacasgsc 2361 GCUGUGAGAGAGAUGCUCAAUAU 2700AD-1730077 asgsgcuaUfgAfCfAfaagucaagguL96 1659asCfscudTg(Agn)cuuuguCfaUfagccusgsg 2362 CCAGGCUAUGACAAAGUCAAGGA 2705AD-1730103 ususccuuUfgUfAfCfuggaggaguuL96 1660asAfscudCc(Tgn)ccaguaCfaAfaggaascsc 2363 GGUUCCUUUGUACUGGAGGAGUG 2711AD-1730108 ususguacUfgGfAfGfgagugagucuL96 1555asGfsacdTc(Agn)cuccucCfaGfuacaasasg 2364 CUUUGUACUGGAGGAGUGAGUCC 2713AD-1730110 gsusacugGfaGfGfAfgugagucccuL96 1661asGfsggdAc(Tgn)cacuccUfcCfaguacsasa 2365 UUGUACUGGAGGAGUGAGUCCCU 52715AD-1730112 ascsuggaGfgAfGfUfgagucccuauL96 1662asUfsagdGg(Agn)cucacuCfcUfccagusasc 2366 GUACUGGAGGAGUGAGUCCCUAU 2716AD-1730118 gsgsagugAfgUfCfCfcuaugcugauL96 1663asUfscadGc(Agn)uagggaCfuCfacuccsusc 2367 GAGGAGUGAGUCCCUAUGCUGAC 2719AD-1730122 asasuacuUfgCfAfGfaggugauucuL96 1556asGfsaadTc(Agn)ccucugCfaAfguauusgsg 2368 CCAAUACUUGCAGAGGUGAUUCU 2722AD-1730133 usgsauagUfuCfAfCfaagagaaguuL96 1558asAfscudTc(Tgn)cuugugAfaCfuaucasasg 2369 CUUGAUAGUUCACAAGAGAAGUC 2726AD-1730143 csasagagAfaGfUfCfguuucauucuL96 1568asGfsaadTg(Agn)aacgacUfuCfucuugsusg 2370 CACAAGAGAAGUCGUUUCAUUCA 2736AD-1730164 gsasguagUfgGfAfUfgucugcaaauL96 1575asUfsuudGc(Agn)gacaucCfaCfuacucscsc 2371 GGGAGUAGUGGAUGUCUGCAAAA 2744AD-1730167 usasguggAfuGfUfCfugcaaaaacuL96 1578asGfsuudTu(Tgn)gcagacAfuCfcacuascsu 2372 AGUAGUGGAUGUCUGCAAAAACC 2747AD-1730168 asgsuggaUfgUfCfUfgcaaaaaccuL96 1664asGfsgudTu(Tgn)ugcagaCfaUfccacusasc 2373 GUAGUGGAUGUCUGCAAAAACCA 2748AD-1730169 gsusggauGfuCfUfGfcaaaaaccauL96 1665asUfsggdTu(Tgn)uugcagAfcAfuccacsusa 2374 UAGUGGAUGUCUGCAAAAACCAG 2749AD-1730171 gsgsauguCfuGfCfAfaaaaccagauL96 1579asUfscudGg(Tgn)uuuugcAfgAfcauccsasc 2375 GUGGAUGUCUGCAAAAACCAGAA 2751AD-1730183 asasaccaGfaAfGfCfggcaaaagcuL96 1666asGfscudTu(Tgn)gccgcuUfcUfgguuususu 2376 AAAAACCAGAAGCGGCAAAAGCA 2759AD-1730184 asasccagAfaGfCfGfgcaaaagcauL96 1585asUfsgcdTu(Tgn)ugccgcUfuCfugguususu 2377 AAAACCAGAAGCGGCAAAAGCAG 2760AD-1730256 usgsgcugAfaGfGfAfgaaacuccauL96 1667asUfsggdAg(Tgn)uucuccUfuCfagccasgsg 2378 CCUGGCUGAAGGAGAAACUCCAA 2764AD-1730287 usgsggauUfgAfAfUfuaaaacagcuL96 1592asGfscudGu(Tgn)uuaauuCfaAfucccascsg 2379 CGUGGGAUUGAAUUAAAACAGCU 2770AD-1730288 gsgsgauuGfaAfUfUfaaaacagcuuL96 1593asAfsgcdTg(Tgn)uuuaauUfcAfaucccsasc 2380 GUGGGAUUGAAUUAAAACAGCUG 2771AD-1730293 usgsaauuAfaAfAfCfagcugcgacuL96 1668asGfsucdGc(Agn)gcuguuUfuAfauucasasu 2381 AUUGAAUUAAAACAGCUGCGACA 2774AD-1730476 asasuuaaAfaCfAfGfcugcgacaauL96 1669asUfsuguCfgcagcugUfuUfuaauuscsa 2382 UGAAUUAAAACAGCUGCGACAAC 2776AD-1730477 asasuuaaaaCfAfGfcugcgacaauL96 1670asdTsugdTcdGcagedTgUfuuuaauuscsa 2383 UGAAUUAAAACAGCUGCGACAAC 2776AD-1730478 asusuaaaacAfGfCfugcgacaacuL96 1671asdGsuudGudCgcagdCuGfuuuuaaususc 2384 GAAUUAAAACAGCUGCGACAACA 2777

Example 2. In Vitro Screening Methods Cell Culture and Transfections:

Transfection assays were carried out in primary human hepatocytes (PHH,BioIVT). Transfection was performed by adding of 5 μl Opti-MEM plus 0.1μl of Lipofectamine RNAiMax per well (Invitrogen, Carlsbad Calif. cat#13778-150) to 5 μl of siRNA duplexes per well into a 384-well plate andincubated at room temperature for 15 minutes. 40 μl of in Invitrogro CPmedia (BioIVT, Cat #Z99029) containing ˜10×10³ cells were then added tothe siRNA mixture. Cells were incubated for 24 hours prior to RNApurification. Experiments were performed at 10 nM, 1 nM, and 0.1 nM.

Total RNA Isolation Using DYNABEADS mRNA Isolation Kit:

RNA was isolated using an Highres Biosolution integration system usingDynabeads™ mRNA DIRECT™ Purification Kit (Invitrogen™, Catalog No.61012). Briefly, 70 μL of Lysis/Binding Buffer and 10 μL of lysis buffercontaining 3 μL of magnetic beads were added to the plate with cells.Plates were incubated on an electromagnetic shaker for 10 minutes atroom temperature and then magnetic beads were captured and thesupernatant was removed. Bead-bound RNA was then washed 2 times with 90μL Wash Buffer A and once with 90 μL Wash Buffer B. Beads were thenwashed with 90 μL Elution Buffer, re-captured, and supernatant wasremoved. Complementary DNA (cDNA) was synthesized using High-CapacitycDNA Reverse Transcription Kit with RNase Inhibitor (AppliedBiosystems™, Catalog No. 4374967) according to the manufacturer'srecommendations. A master mix containing 1 μL 10× Buffer, 0.4 μL 25×deoxyribonucleotide triphosphate, 1 μL 10× Random primers, 0.5 μLReverse Transcriptase, 0.5 μL RNase inhibitor, and 6.6 μL of H₂O perreaction was added to RNA isolated above. The plates were sealed, mixed,and incubated on an electromagnetic shaker for 10 minutes at roomtemperature, followed by 2 hours incubation at 37° C.

CFB mRNA levels were quantified by performing RT-qPCR analysis. 2 μl ofcDNA were added to a master mix containing 0.5 μl of human or cyno GAPDHTaqMan Probe, 0.5 μl human or cyno CFB probe (Hs00156060_m1) and 5 μlLightcycler 480 probe master mix (Roche Cat #04887301001) per well in a384 well plates. Real time PCR was done in a LightCycler480 Real TimePCR system (Roche). To calculate relative fold change, real-time datawere analyzed using the Delta-Delta Threshold Cycle (RelativeQuantification) (ΔΔC_(q)[RQ]) method [Schmittgen and Livak 2008] andwere normalized to control assays performed using cells transfected withPBS. For all samples, CFB mRNA levels were first normalized to GAPDH asa reference gene. Data are expressed as percent of CFB mRNA remainingrelative to average PBS control and error is expressed as standarddeviation (SD), derived from the 4 transfection replicates.

The results of single dose transfection screens in PHH cells of thedsRNA agents in Tables 2 and 3 are shown in Table 4.

TABLE 4 In Vitro Single dose Screens in Primary Human Hepatocytes 10 nM1 nM 0.1 nM Average Average Average mRNA mRNA mRNA remaining 10 nMremaining 1 nM remaining 0.1 nM Duplex Name (%) STDEV (%) STDEV (%)STDEV AD-1728447.1 14 4 13 4 21 3 AD-1724362.1 17 10 13 3 20 4AD-1724363.1 19 6 26 7 25 1 AD-1724364.1 16 5 19 1 15 4 AD-1724365.1 3813 47 8 36 4 AD-1724369.1 23 7 27 7 30 9 AD-1724370.1 25 7 48 5 48 8AD-1724376.1 20 4 19 3 23 6 AD-1728461.1 22 5 28 3 37 5 AD-1724384.1 697 52 9 78 14 AD-1728470.1 42 11 43 10 71 12 AD-1724385.1 47 7 66 6 53 9AD-1728471.1 30 1 40 7 37 10 AD-1724386.1 37 3 49 6 69 15 AD-1724530.114 1 30 6 30 5 AD-1724572.1 31 5 62 8 75 18 AD-1728659.1 26 5 36 6 45 9AD-1724574.1 13 3 16 0 16 4 AD-1724575.1 17 1 32 5 31 6 AD-1724576.1 111 15 1 18 3 AD-1724579.1 21 2 21 2 34 7 AD-1728664.1 76 2 85 10 95 9AD-1726815.1 55 5 67 6 79 9 AD-1728671.1 55 5 60 9 84 8 AD-1724586.1 272 41 8 52 4 AD-1728685.1 60 8 66 8 76 11 AD-1724600.1 21 4 32 5 34 5AD-1728736.1 12 2 17 4 13 2 AD-1724651.1 8 1 11 1 10 2 AD-1724653.1 13 217 3 21 6 AD-1724685.1 15 2 33 2 27 3 AD-1726927.1 12 2 22 3 29 5AD-1724691.1 13 1 17 2 21 2 AD-1728777.1 25 8 50 7 44 6 AD-1726928.1 203 32 2 39 4 AD-1724692.1 9 1 16 2 14 3 AD-1724693.1 8 1 11 1 14 2AD-1726931.1 56 5 61 5 75 3 AD-1724695.1 12 2 24 4 21 1 AD-1726934.1 253 28 10 49 6 AD-1724698.1 18 2 21 1 25 2 AD-1728784.1 7 1 12 2 10 1AD-1726935.1 9 2 10 2 13 2 AD-1724699.1 6 1 9 1 6 1 AD-1726936.1 6 2 102 12 3 AD-1724700.1 8 1 13 2 10 2 AD-1728786.1 9 1 14 4 12 2AD-1726937.1 7 1 10 1 9 2 AD-1724701.1 9 2 12 1 8 1 AD-1728787.1 54 8 627 69 11 AD-1726938.1 12 3 17 3 18 5 AD-1724702.1 6 1 12 3 9 2AD-1726939.1 4 1 8 1 9 1 AD-1724703.1 7 1 12 3 12 2 AD-1728789.1 12 2 193 22 5 AD-1726940.1 10 1 14 4 15 2 AD-1724704.1 8 1 11 2 10 1AD-1726941.1 14 1 23 8 27 6 AD-1724705.1 13 2 19 6 20 4 AD-1724706.1 3 110 2 9 1 AD-1726942.1 7 1 11 3 10 2 AD-1724707.1 10 1 25 2 20 3AD-1728793.1 14 2 21 8 22 5 AD-1724708.1 10 1 11 1 12 1 AD-1724714.1 122 19 5 25 5 AD-1724715.1 10 2 16 5 14 3 AD-1728801.1 6 1 15 8 38 2AD-1726952.1 12 4 10 3 34 9 AD-1724716.1 15 2 18 3 26 5 AD-1728802.1 529 79 13 86 16 AD-1724717.1 31 7 45 9 74 23 AD-1724718.1 8 1 14 3 16 2AD-1726961.1 6 0 20 3 20 3 AD-1724725.1 5 3 15 2 17 2 AD-1728810.1 59 1546 11 46 17 AD-1728811.1 35 12 46 4 58 11 AD-1724726.1 20 8 36 7 40 15AD-1724730.1 16 4 23 7 27 5 AD-1724731.1 3 9 53 10 77 16 AD-1724741.1 428 52 9 64 23 AD-1724742.1 14 4 22 7 22 5 AD-1728827.1 31 10 43 3 51 8AD-1724743.1 23 10 23 5 60 18 AD-1728861.1 28 10 20 9 28 12 AD-1727012.115 5 18 4 22 8 AD-1724776.1 11 4 15 4 18 5 AD-1724777.1 12 6 18 6 25 9AD-1728863.1 15 5 21 6 33 13 AD-1724778.1 13 3 20 4 24 5 AD-1724779.1 114 26 5 37 5 AD-1724780.1 47 6 65 10 88 13 AD-1724781.1 19 4 32 6 42 3AD-1728877.1 23 4 32 8 37 7 AD-1724792.1 29 9 38 3 47 7 AD-1724819.1 101 15 4 21 4 AD-1724823.1 12 1 13 3 23 9 AD-1727059.1 12 2 18 2 15 2AD-1728909.1 23 5 3 11 46 5 AD-1724824.1 21 4 35 2 30 4 AD-1724825.1 224 27 15 44 7 AD-1724860.1 51 11 55 13 73 6 AD-1724894.1 17 3 31 6 41 9AD-1724897.1 16 1 27 5 26 4 AD-1724899.1 13 4 21 6 19 2 AD-1724900.1 112 22 2 26 3 AD-1724903.1 44 5 46 9 69 14 AD-1727140.1 34 21 47 12 75 10AD-1724904.1 40 4 54 12 54 11 AD-1728990.1 13 2 22 3 16 2 AD-1724905.115 1 22 3 25 4 AD-1727142.1 51 11 77 12 81 8 AD-1724906.1 42 6 54 5 53 2AD-1728995.1 25 4 37 1 26 7 AD-1724910.1 14 1 20 3 12 1 AD-1724919.1 255 46 9 38 4 AD-1727181.1 52 11 71 19 76 4 AD-1724945.1 17 4 23 6 26 6AD-1724946.1 31 6 32 6 61 4 AD-1727183.1 17 4 14 7 20 2 AD-1724947.1 156 8 2 17 4 AD-1727184.1 84 9 87 3 94 11 AD-1724948.1 37 8 43 5 52 5AD-1724949.1 35 1 52 11 61 8 AD-1725000.1 27 3 47 9 36 5 AD-1725003.1 5916 65 9 65 5 AD-1729089.1 112 19 122 19 90 20 AD-1725004.1 97 16 106 887 13 AD-1725013.1 27 1 43 5 39 5 AD-1725017.1 48 4 75 11 82 9AD-1729103.1 52 3 74 4 78 9 AD-1725018.1 39 4 43 3 56 7 AD-1725019.1 717 102 8 90 7 AD-1729105.1 66 9 84 17 71 13 AD-1725020.1 18 3 31 5 24 2AD-1729106.1 32 4 48 8 48 6 AD-1725021.1 19 1 30 6 24 3 AD-1725022.1 356 43 7 43 10 AD-1725023.1 23 1 39 5 40 2 AD-1729110.1 23 6 36 4 32 4AD-1725025.1 16 5 29 5 26 6 AD-1729112.1 35 6 49 11 46 8 AD-1727263.1 364 52 5 68 13 AD-1725027.1 35 7 47 3 41 11 AD-1725028.1 21 4 44 7 42 3AD-1725033.1 73 11 89 15 88 10 AD-1727275.1 52 6 52 8 41 2 AD-1725039.126 4 28 3 20 2 AD-1727276.1 107 18 99 11 101 10 AD-1725040.1 29 3 41 241 13 AD-1725041.1 13 3 24 5 17 3 AD-1727278.1 76 16 83 16 86 10AD-1725042.1 17 3 24 6 21 3 AD-1725043.1 12 5 21 3 18 3 AD-1725044.1 7 314 5 11 1 AD-1729130.1 62 13 81 13 69 7 AD-1725045.1 16 3 25 3 17 4AD-1725046.1 19 2 21 5 23 6 AD-1729132.1 62 9 50 6 39 6 AD-1725047.1 191 25 5 17 3 AD-1725048.1 15 1 19 2 17 4 AD-1729134.1 34 3 55 11 56 8AD-1727285.1 23 5 35 3 44 11 AD-1725049.1 14 2 21 5 19 5 AD-1727286.1 324 49 9 50 6 AD-1725050.1 17 4 31 9 27 7 AD-1729136.1 16 3 22 3 20 9AD-1725051.1 10 0 23 3 17 1 AD-1729137.1 7 0 11 2 9 1 AD-1727288.1 5 213 2 8 2 AD-1725052.1 9 3 12 1 9 4 AD-1727289.1 9 2 17 2 18 9AD-1725053.1 11 2 20 4 17 3 AD-1729139.1 7 1 12 2 9 1 AD-1727290.1 8 115 4 12 5 AD-1725054.1 5 2 5 0 6 1 AD-1727291.1 9 1 8 2 14 3AD-1725055.1 6 2 6 2 6 0 AD-1729141.1 9 3 10 4 12 1 AD-1727292.1 7 3 6 215 1 AD-1725056.1 15 4 13 2 17 4 AD-1725057.1 8 1 7 2 5 2 AD-1729142.116 1 19 5 23 5 AD-1727293.1 9 1 7 1 5 1 AD-1725058.1 12 3 11 4 25 4AD-1725059.1 8 3 5 1 12 1 AD-1725060.1 22 2 13 5 9 1 AD-1725061.1 18 413 3 9 1 AD-1727298.1 29 3 21 3 11 1 AD-1725062.1 27 3 19 2 8 1AD-1725066.1 28 5 15 2 14 5 AD-1729151.1 40 4 27 3 22 4 AD-1727310.1 856 57 13 71 6 AD-1725074.1 25 1 16 2 11 1 AD-1725075.1 22 1 13 3 11 1AD-1725079.1 38 8 22 4 21 3 AD-1725080.1 65 4 46 5 59 2 AD-1727318.1 383 25 3 15 1 AD-1725082.1 26 6 15 4 8 1 AD-1725083.1 30 7 19 5 11 2AD-1725088.1 69 5 41 10 40 4 AD-1725092.1 47 7 31 8 1 2 AD-1725095.1 7 26 2 5 1 AD-1729180.1 52 5 54 7 57 9 AD-1727331.1 28 3 21 6 30 3AD-1725096.1 18 1 11 3 6 1 AD-1729207.1 76 9 59 8 90 11 AD-1727358.1 424 38 7 50 6 AD-1725122.1 31 3 21 4 21 2 AD-1727359.1 62 11 38 4 29 5AD-1725123.1 33 3 22 2 41 8 AD-1725125.1 21 5 11 3 6 1 AD-1727361.1 26 315 5 9 2 AD-1727392.1 24 2 13 2 9 1 AD-1725156.1 19 1 11 5 16 3AD-1729242.1 22 2 15 6 11 1 AD-1725158.1 50 6 32 6 40 8 AD-1725159.1 6310 45 8 42 6 AD-1729269.1 52 1 51 5 51 3 AD-1727420.1 24 4 18 1 13 3AD-1725184.1 22 4 16 2 9 1 AD-1729271.1 47 5 31 5 23 3 AD-1725186.1 39 521 1 16 2 AD-1729274.1 43 5 37 2 40 5 AD-1725189.1 29 5 17 2 14 1AD-1725190.1 22 5 14 5 12 1 AD-1727427.1 74 8 56 5 66 8 AD-1725191.1 273 25 4 21 1 AD-1729277.1 51 12 33 5 35 2 AD-1727428.1 97 4 65 9 70 4AD-1725192.1 20 2 15 2 9 1 AD-1725193.1 18 1 9 1 12 4 AD-1727430.1 26 616 4 17 2 AD-1725194.1 13 2 9 1 8 1 AD-1729280.1 27 5 23 2 20 5AD-1727431.1 30 8 21 4 20 2 AD-1725195.1 25 6 16 3 13 3 AD-1727432.1 194 14 4 9 1 AD-1725196.1 18 5 15 1 11 1 AD-1727433.1 68 19 43 7 65 4AD-1725197.1 33 12 27 5 52 3 AD-1727434.1 50 10 49 3 36 2 AD-1725198.124 5 21 2 23 3 AD-1727435.1 46 13 36 4 35 4 AD-1725199.1 30 7 27 3 26 6AD-1729285.1 37 8 38 7 32 3 AD-1727436.1 36 8 25 3 36 9 AD-1725200.1 175 16 2 14 1 AD-1729288.1 98 12 91 17 92 15 AD-1725203.1 64 16 69 8 52 4AD-1725204.1 77 18 77 17 73 7 AD-1729290.1 26 4 25 2 5 2 AD-1727441.1 4910 52 7 55 7 AD-1725205.1 31 7 35 1 27 4 AD-1725206.1 23 4 31 4 39 4AD-1727442.1 45 11 44 6 48 5 AD-1725208.1 13 4 16 2 15 2 AD-1729296.1 288 33 6 16 5 AD-1725211.1 28 9 32 3 39 3 AD-1725212.1 16 9 21 3 20 4AD-1729300.1 58 16 48 6 77 15 AD-1725215.1 15 3 20 6 33 5 AD-1725216.120 6 28 12 33 5 AD-1725243.1 32 4 37 10 49 8 AD-1725244.1 12 3 16 4 10 3AD-1727481.1 22 2 36 4 23 0 AD-1725245.1 16 1 22 3 15 4 AD-1727483.1 302 41 7 48 9 AD-1725247.1 13 2 25 3 24 3 AD-1725327.1 22 4 35 7 27 5AD-1729413.1 68 19 82 10 87 6 AD-1725328.1 39 3 57 9 75 10 AD-1727565.17 6 90 5 94 5 AD-1725329.1 54 8 91 15 103 16 AD-1727566.1 70 10 90 9 10513 AD-1725330.1 62 10 76 6 92 10 AD-1725331.1 31 4 67 4 73 17AD-1727568.1 85 10 105 11 88 8 AD-1725332.1 54 5 59 8 63 7 AD-1727569.121 3 37 7 41 7 AD-1725333.1 24 4 38 4 44 6 AD-1727570.1 22 4 34 5 38 5AD-1725334.1 20 4 32 4 26 8 AD-1725336.1 17 5 34 10 30 6 AD-1725344.1 396 54 10 64 17 AD-1725345.1 38 2 62 5 68 5 AD-1725347.1 35 3 50 11 49 4AD-1727584.1 32 5 44 2 36 7 AD-1725348.1 15 3 24 1 22 4 AD-1727612.1 358 20 1 40 5 AD-1725376.1 16 6 45 13 41 7 AD-1729461.1 66 13 78 16 79 3AD-1729462.1 34 19 93 6 99 8 AD-1725377.1 62 15 87 16 89 9 AD-1729463.128 6 58 2 46 5 AD-1725378.1 25 4 44 4 43 5 AD-1727633.1 47 7 64 14 58 11AD-1725397.1 16 3 24 6 20 5 AD-1729487.1 13 0 21 1 10 3 AD-1727638.1 111 23 2 15 3 AD-1725402.1 14 2 25 3 15 3 AD-1727639.1 14 2 23 6 14 3AD-1725403.1 12 3 21 4 13 2 AD-1727640.1 37 5 60 6 68 11 AD-1725404.1 162 35 3 27 7 AD-1727641.1 13 2 26 5 20 1 AD-1725405.1 9 1 15 2 11 2AD-1727642.1 47 8 82 13 78 8 AD-1725406.1 17 1 26 4 23 4 AD-1727643.1 211 36 9 31 5 AD-1725407.1 13 1 26 3 20 5 AD-1727644.1 14 1 24 2 23 4AD-1725408.1 8 1 15 1 10 1 AD-1727645.1 14 2 21 2 15 2 AD-1725409.1 12 316 3 12 2 AD-1727646.1 47 6 74 10 83 11 AD-1725410.1 22 1 37 4 32 4AD-1725411.1 24 3 44 6 44 7 AD-1730469.1 72 3 96 1 102 4 AD-1730470.1 364 55 11 76 4 AD-1730471.1 16 3 27 5 30 6 AD-1727663.1 45 4 60 15 54 5AD-1725427.1 18 2 34 2 27 2 AD-1727664.1 32 5 53 3 45 6 AD-1725428.1 213 35 5 29 3 AD-1729514.1 13 2 21 3 15 3 AD-1727665.1 12 3 19 4 20 3AD-1725429.1 11 2 22 5 14 2 AD-1729515.1 37 5 36 5 34 3 AD-1727666.1 152 23 4 14 3 AD-1725430.1 11 1 13 5 13 3 AD-1729524.1 30 3 56 5 49 4AD-1727675.1 24 5 35 9 26 5 AD-1725439.1 29 6 44 10 45 10 AD-1725440.125 1 32 4 29 3 AD-1729525.1 29 8 47 8 59 11 AD-1727677.1 30 6 47 6 41 2AD-1725441.1 21 2 28 1 26 2 AD-1725449.1 32 7 41 7 29 8 AD-1729538.1 293 55 4 51 8 AD-1727689.1 41 5 67 8 61 9 AD-1725453.1 29 9 47 5 54 8AD-1727690.1 9 4 14 5 16 4 AD-1725454.1 15 3 18 4 24 2 AD-1729541.1 9 426 2 17 4 AD-1725456.1 13 3 18 1 15 2 AD-1727693.1 11 1 18 3 11 2AD-1725457.1 9 2 20 2 11 3 AD-1729545.1 15 3 22 4 18 2 AD-1727696.1 12 39 3 15 3 AD-1725460.1 5 2 8 2 11 2 AD-1727698.1 19 1 20 2 23 10AD-1725462.1 5 1 9 3 8 3 AD-1729548.1 7 2 9 2 7 3 AD-1727699.1 7 2 7 210 2 AD-1725463.1 4 1 7 3 7 1 AD-1725464.1 3 1 8 1 10 1 AD-1729550.1 261 24 11 38 4 AD-1727701.1 13 2 28 2 33 5 AD-1725465.1 9 1 19 4 19 3AD-1729552.1 25 3 33 3 26 4 AD-1727703.1 6 1 11 2 10 1 AD-1725467.1 7 112 2 14 3 AD-1725469.1 7 2 11 1 16 2 AD-1727705.1 16 2 29 2 29 8AD-1729555.1 7 1 10 1 13 1 AD-1725470.1 11 1 17 2 19 5 AD-1729557.1 42 660 3 48 4 AD-1727708.1 12 3 16 2 19 3 AD-1725472.1 9 0 13 2 11 2AD-1725473.1 11 2 18 3 18 1 AD-1729559.1 18 4 28 6 45 4 AD-1727710.1 396 68 11 70 7 AD-1725474.1 19 4 40 2 38 3 AD-1729561.1 11 1 17 1 22 3AD-1727712.1 10 1 16 1 19 1 AD-1725476.1 6 1 9 1 13 1 AD-1729562.1 4 3 41 6 2 AD-1727713.1 8 1 10 1 12 1 AD-1725477.1 7 0 10 0 13 1 AD-1727714.19 2 15 1 20 3 AD-1725478.1 9 1 12 1 12 1 AD-1727717.1 23 2 38 4 53 6AD-1725481.1 17 2 29 6 37 1 AD-1727718.1 19 2 33 3 40 3 AD-1725482.1 163 31 4 31 9 AD-1729567.1 65 3 60 8 70 6 AD-1729568.1 20 4 23 3 25 4AD-1725483.1 13 1 12 2 16 1 AD-1725534.1 13 2 23 8 38 8 AD-1725535.1 6 16 1 9 0 AD-1725548.1 5 2 12 3 19 3 AD-1725552.1 29 6 67 7 75 9AD-1725556.1 13 6 22 6 29 5 AD-1729643.1 9 6 24 6 26 5 AD-1725558.1 20 332 8 40 6 AD-1725580.1 25 5 32 5 35 4 AD-1729667.1 43 7 59 4 68 10AD-1725582.1 46 8 47 7 55 9 AD-1729670.1 12 3 25 4 34 3 AD-1727821.1 192 38 9 46 6 AD-1725585.1 15 1 27 4 41 6 AD-1727823.1 72 8 71 11 77 10AD-1725587.1 21 2 43 1 48 11 AD-1729673.1 22 2 32 8 33 7 AD-1725588.1 194 20 4 31 5 AD-1725590.1 17 4 36 4 55 8 AD-1727826.1 99 18 83 7 84 15AD-1725591.1 42 5 77 4 78 14 AD-1729677.1 17 2 24 1 29 6 AD-1725592.1 202 34 2 47 4 AD-1727829.1 88 22 89 14 93 7 AD-1725593.1 14 2 17 3 33 7AD-1725598.1 94 24 66 11 79 18 AD-1729688.1 24 3 54 12 61 13AD-1725603.1 11 3 20 3 23 2 AD-1725604.1 15 4 17 3 29 4 AD-1729690.1 889 83 12 86 17 AD-1725605.1 64 11 49 8 67 12 AD-1725643.1 7 0 10 2 15 1AD-1725644.1 2 0 11 3 13 3 AD-1729729.1 28 6 31 7 41 11 AD-1725645.1 187 42 7 41 5 AD-1725646.1 13 5 37 7 46 9 AD-1727883.1 16 6 28 6 44 9AD-1725647.1 14 3 17 5 29 8 AD-1725667.1 23 4 30 6 40 6 AD-1725716.1 6714 49 12 62 15 AD-1725717.1 22 2 46 9 63 14 AD-1729802.1 102 26 77 11 7913 AD-1729841.1 3 1 7 2 11 2 AD-1727977.1 2 0 6 1 12 5 AD-1725756.1 3 06 2 10 2 AD-1727978.1 11 2 11 2 15 4 AD-1725757.1 6 1 7 2 10 2AD-1727980.1 7 2 8 2 11 4 AD-1725759.1 7 2 6 1 13 2 AD-1727981.1 6 0 7 110 1 AD-1725760.1 7 1 8 2 11 1 AD-1725761.1 6 2 7 2 9 2 AD-1725762.1 9 317 2 19 3 AD-1727984.1 7 1 9 1 10 2 AD-1725763.1 3 1 5 1 8 2AD-1727985.1 7 1 10 2 17 4 AD-1725764.1 4 1 5 2 11 1 AD-1729849.1 42 1031 5 41 11 AD-1729850.1 12 4 14 3 21 2 AD-1727986.1 5 1 9 2 13 1AD-1725765.1 8 1 14 2 22 2 AD-1727987.1 6 1 8 1 12 2 AD-1725766.1 5 1 92 12 2 AD-1725767.1 6 1 10 2 14 1 AD-1729852.1 33 7 43 7 58 16AD-1727989.1 10 2 13 3 16 2 AD-1725768.1 9 2 15 3 18 2 AD-1729854.1 21 131 5 32 3 AD-1727990.1 10 1 17 2 19 3 AD-1725769.1 8 1 14 1 20 2AD-1727992.1 9 1 15 4 21 2 AD-1725771.1 8 2 12 3 7 4 AD-1729856.1 14 424 3 29 2 AD-1727993.1 13 1 24 3 18 6 AD-1725772.1 7 1 9 1 10 2AD-1727994.1 23 2 40 6 43 7 AD-1725773.1 7 2 11 3 15 1 AD-1727996.1 19 830 2 53 7 AD-1725775.1 45 10 55 8 71 8 AD-1729861.1 60 5 69 2 79 6AD-1725776.1 23 6 29 9 57 9 AD-1729862.1 37 10 42 6 42 2 AD-1725777.1 82 12 3 15 2 AD-1727999.1 50 7 64 14 78 9 AD-1725778.1 17 5 25 3 35 3AD-1725779.1 16 1 21 1 29 4 AD-1725780.1 7 1 11 1 11 2 AD-1729869.1 28 143 5 57 11 AD-1725784.1 10 2 15 1 17 1 AD-1729870.1 30 2 50 13 59 4AD-1725785.1 13 1 21 3 34 2 AD-1725786.1 16 4 22 3 30 2 AD-1729872.1 443 51 12 54 5 AD-1725787.1 16 3 24 3 28 5 AD-1725789.1 11 1 16 2 20 3AD-1725790.1 20 1 29 4 37 6 AD-1728049.1 102 14 94 11 100 14AD-1725828.1 26 5 34 6 46 9 AD-1725829.1 44 7 50 8 64 3 AD-1725830.1 112 21 5 26 5 AD-1725831.1 18 5 21 3 24 2 AD-1725832.1 15 1 26 4 37 4AD-1728061.1 38 5 40 5 50 10 AD-1725840.1 43 3 32 4 29 2 AD-1729926.1 446 50 4 45 5 AD-1728062.1 38 4 37 4 46 6 AD-1725841.1 13 2 15 2 20 3AD-1725842.1 12 2 17 2 22 6 AD-1725845.1 56 12 74 7 81 18 AD-1728067.185 10 98 21 108 12 AD-1725846.1 42 6 68 7 75 11 AD-1729933.1 16 1 22 521 1 AD-1725848.1 12 2 14 1 15 3 AD-1725849.1 14 4 17 5 28 5AD-1725850.1 31 4 39 9 58 6 AD-1725854.1 12 1 15 7 23 7 AD-1725855.1 8 110 5 19 2 AD-1729941.1 30 3 47 7 59 5 AD-1725856.1 20 4 32 1 39 6AD-1725857.1 10 0 15 2 18 4 AD-1725858.1 25 4 36 8 46 7 AD-1725861.1 396 41 7 66 12 AD-1729947.1 51 7 46 14 53 2 AD-1725862.1 40 4 51 6 66 7AD-1728085.1 25 5 50 10 48 8 AD-1725864.1 11 2 20 3 25 4 AD-1729951.1 277 45 7 59 11 AD-1725866.1 16 4 29 7 40 9 AD-1725867.1 24 6 34 6 54 5AD-1725872.1 15 2 17 3 26 6 AD-1725874.1 20 8 25 3 39 8 AD-1729992.1 608 34 8 41 13 AD-1725907.1 45 10 29 10 54 10 AD-1729993.1 51 9 25 4 47 13AD-1725908.1 15 0 13 4 19 2 AD-1729994.1 47 12 60 10 79 18 AD-1725909.117 3 29 10 42 7 AD-1728132.1 31 7 35 7 65 4 AD-1725911.1 36 5 36 7 58 10AD-1730001.1 6 1 10 4 21 4 AD-1728137.1 6 1 10 2 19 3 AD-1725916.1 5 110 2 12 1 AD-1728140.1 7 1 16 6 28 4 AD-1725919.1 7 3 16 5 22 5AD-1728146.1 10 1 16 2 26 8 AD-1725925.1 6 1 10 2 24 7 AD-1730042.1 18 237 4 65 14 AD-1725957.1 24 3 32 3 50 9 AD-1725958.1 18 1 28 6 42 8AD-1725961.1 8 1 16 4 21 3 AD-1730048.1 10 1 17 2 27 5 AD-1725963.1 15 319 4 26 4 AD-1725964.1 18 4 15 3 22 6 AD-1725967.1 15 3 26 8 33 4AD-1730053.1 62 7 51 11 73 17 AD-1725968.1 46 7 39 7 56 7 AD-1730059.131 6 51 6 85 23 AD-1728195.1 23 4 38 4 67 13 AD-1725974.1 11 3 16 1 4613 AD-1725977.1 20 4 24 2 25 15 AD-1730068.1 11 1 13 2 11 5 AD-1728204.115 1 18 3 38 5 AD-1725983.1 11 2 15 3 24 4 AD-1728206.1 11 1 16 3 40 10AD-1725985.1 7 2 12 2 21 2 AD-1728207.1 12 3 23 2 25 7 AD-1725986.1 18 422 3 25 4 AD-1728208.1 13 1 22 1 35 2 AD-1725987.1 13 3 17 2 24 2AD-1728209.1 46 6 70 6 101 10 AD-1725988.1 20 7 28 5 43 7 AD-1725989.123 4 38 1 66 7 AD-1728210.1 26 4 42 1 65 8 AD-1728212.1 16 2 23 3 57 7AD-1725991.1 10 1 18 4 31 6 AD-1730077.1 46 2 62 6 102 8 AD-1725992.1 332 54 8 92 10 AD-1728214.1 17 2 30 5 74 16 AD-1725993.1 13 2 25 7 50 5AD-1728220.1 10 2 10 4 34 2 AD-1725999.1 11 2 16 3 32 3 AD-1726014.1 415 68 13 100 31 AD-1726015.1 30 5 53 11 98 16 AD-1726016.1 49 9 76 14 1145 AD-1730103.1 38 8 56 7 92 12 AD-1726018.1 29 10 42 4 80 10AD-1726020.1 34 5 45 3 74 24 AD-1730108.1 63 5 79 13 109 14 AD-1728244.158 9 85 9 109 12 AD-1726023.1 40 6 44 7 105 14 AD-1726024.1 31 9 51 9 8818 AD-1730110.1 44 6 53 4 108 17 AD-1726025.1 21 3 34 9 65 10AD-1730112.1 62 12 76 9 92 10 AD-1726027.1 23 5 36 4 66 14 AD-1726029.117 5 41 7 76 7 AD-1726031.1 20 3 31 5 53 10 AD-1730118.1 48 8 74 13 9613 AD-1726033.1 34 4 64 5 99 11 AD-1726034.1 18 5 33 5 53 11AD-1726036.1 7 1 15 4 22 2 AD-1728258.1 3 0 15 4 43 3 AD-1726037.1 9 215 4 32 7 AD-1730122.1 12 3 18 3 37 6 AD-1728260.1 21 2 34 4 70 7AD-1726039.1 15 1 22 3 55 12 AD-1726041.1 19 2 35 5 75 13 AD-1726042.136 8 52 11 96 13 AD-1730472.1 14 5 24 7 66 11 AD-1730133.1 8 2 21 2 32 4AD-1728269.1 12 2 25 2 55 10 AD-1726048.1 7 2 13 1 24 1 AD-1728270.1 145 35 8 90 20 AD-1726049.1 18 6 42 7 81 6 AD-1728271.1 11 4 20 2 38 10AD-1726050.1 8 2 13 3 19 4 AD-1726051.1 4 1 6 2 10 3 AD-1728272.1 20 430 3 64 16 AD-1728273.1 4 2 9 4 20 3 AD-1726052.1 4 1 9 2 19 5AD-1728274.1 7 2 13 1 28 7 AD-1726053.1 5 1 9 3 17 4 AD-1728275.1 8 4 154 35 9 AD-1726054.1 6 3 12 3 22 6 AD-1728276.1 4 1 8 4 15 5 AD-1726055.16 2 11 2 15 1 AD-1728277.1 7 1 9 2 17 3 AD-1726056.1 6 1 7 1 13 2AD-1728278.1 5 1 10 3 12 3 AD-1726057.1 6 1 7 1 14 2 AD-1726058.1 7 1 123 11 5 AD-1730143.1 8 2 14 1 19 3 AD-1728279.1 7 1 13 2 21 2AD-1726059.1 8 1 16 2 18 3 AD-1726060.1 9 1 16 4 18 2 AD-1728282.1 17 229 9 62 11 AD-1726061.1 9 2 17 2 33 7 AD-1728283.1 15 2 30 5 39 3AD-1726062.1 7 0 14 3 15 2 AD-1728284.1 12 1 20 2 28 5 AD-1726063.1 11 015 3 29 6 AD-1728285.1 8 1 16 3 31 3 AD-1726064.1 11 1 17 5 27 5AD-1728286.1 14 4 25 4 64 10 AD-1726065.1 9 1 14 1 27 6 AD-1730474.1 183 28 11 56 6 AD-1730473.1 15 2 28 5 56 8 AD-1730475.1 9 1 13 2 20 3AD-1730164.1 29 4 42 4 47 7 AD-1728300.1 18 2 27 2 38 1 AD-1726079.1 142 23 7 35 4 AD-1728301.1 12 2 16 2 24 3 AD-1726080.1 9 2 14 2 20 4AD-1728302.1 6 2 7 3 9 2 AD-1726081.1 7 3 9 3 16 3 AD-1730167.1 8 2 9 516 6 AD-1728303.1 9 1 16 1 17 3 AD-1726082.1 10 0 17 2 20 6 AD-1730168.127 6 51 7 73 7 AD-1726083.1 21 3 37 8 63 13 AD-1726084.1 11 1 18 1 24 5AD-1730169.1 20 2 23 3 32 5 AD-1726085.1 9 3 21 2 33 5 AD-1730171.1 21 339 3 49 7 AD-1728307.1 17 1 32 5 50 6 AD-1726086.1 15 5 29 4 53 11AD-1728308.1 16 3 25 3 44 6 AD-1726087.1 11 2 15 2 21 2 AD-1728311.1 5013 73 14 109 15 AD-1726090.1 21 2 42 9 70 10 AD-1728312.1 55 4 60 12 10419 AD-1726091.1 18 3 32 5 46 8 AD-1726092.1 32 5 71 9 82 7 AD-1726095.168 8 73 9 79 12 AD-1728317.1 34 2 40 5 42 9 AD-1726096.1 16 2 24 4 24 5AD-1728318.1 106 19 45 15 110 4 AD-1726097.1 17 4 28 6 38 4 AD-1730183.161 7 71 14 94 16 AD-1726098.1 54 3 69 10 90 9 AD-1730184.1 69 6 74 4 9312 AD-1728320.1 48 9 73 12 94 4 AD-1726099.1 40 7 71 11 79 7AD-1726103.1 70 12 83 11 73 5 AD-1728324.1 67 9 71 12 68 13 AD-1726113.151 7 74 3 88 8 AD-1726159.1 15 4 32 5 39 8 AD-1730256.1 35 6 42 7 56 6AD-1726171.1 15 2 21 2 32 4 AD-1728405.1 30 7 42 5 67 9 AD-1726184.1 113 19 2 30 8 AD-1728408.1 17 1 39 8 63 9 AD-1726187.1 11 2 22 5 39 11AD-1728410.1 9 2 17 3 20 4 AD-1726189.1 7 2 14 1 17 4 AD-1728412.1 9 219 3 29 6 AD-1726191.1 9 1 18 3 26 10 AD-1726201.1 8 1 17 1 16 3AD-1728422.1 12 3 20 6 23 5 AD-1730287.1 13 3 24 2 31 5 AD-1728423.1 153 30 5 37 4 AD-1726202.1 15 2 29 7 44 8 AD-1730288.1 12 1 25 6 41 7AD-1726203.1 11 2 17 4 25 6 AD-1728427.1 16 4 36 4 56 14 AD-1726206.1 122 22 3 28 7 AD-1726207.1 4 1 10 1 11 5 AD-1730293.1 13 4 19 3 23 3AD-1726208.1 6 2 16 2 20 5 AD-1726209.1 7 1 17 4 17 4 AD-1730476.1 22 525 3 17 6 AD-1730477.1 7 1 13 3 13 3 AD-1730478.1 21 3 36 7 50 11

Example 3. In Vitro Screening Methods

A subset of the duplexes was also assessed by transfection and freeuptake in primary human hepatocytes and primary cynomolgus hepatocytes.

Transfection and free uptake assays were carried out in primary humanhepatocyte (PHH, BioIVT) and primary cynomolgus hepatocyte (PCH,BioIVT). Transfection was performed by adding of 5 μl Opti-MEM plus 0.1μl of Lipofectamine RNAiMax per well (Invitrogen, Carlsbad Calif. cat#13778-150) to 5 μl of siRNA duplexes per well into a 384-well plate andincubated at room temperature for 15 minutes. 40 μl of in Invitrogro CPmedia (BioIVT, Cat #Z99029) containing ˜10×10³ cells were then added tothe siRNA mixture. Cells were incubated for 24 hours prior to RNApurification. Experiments were performed at 10 nM, 1 nM, 0.1 nM and 0.01nM. Free uptake assay was performed similarly to transfection assaywithout using Lipofectamine RNAimax and cells were incubated for 48hours prior to the RNA purification. Experiments were performed at 200nM, 100 nM, 10 nM, 1 nM.

RNA was isolated using an Highres Biosolution integration system usingDynabeads™ mRNA DIRECT™ Purification Kit (Invitrogen™, Catalog No.61012). Briefly, 70 μL of Lysis/Binding Buffer and 10 μL of lysis buffercontaining 3 μL of magnetic beads were added to the plate with cells.Plates were incubated on an electromagnetic shaker for 10 minutes atroom temperature and then magnetic beads were captured and thesupernatant was removed. Bead-bound RNA was then washed 2 times with 90μL Wash Buffer A and once with 90 μL Wash Buffer B. Beads were thenwashed with 90 μL Elution Buffer, re-captured, and supernatant wasremoved. Complementary DNA (cDNA) was synthesized using High-CapacitycDNA Reverse Transcription Kit with RNase Inhibitor (AppliedBiosystems™, Catalog No. 4374967) according to the manufacturer'srecommendations. A master mix containing 1 μL 10× Buffer, 0.4 μL 25×deoxyribonucleotide triphosphate, 1 μL 10× Random primers, 0.5 μLReverse Transcriptase, 0.5 μL RNase inhibitor, and 6.6 μL of H₂O perreaction was added to RNA isolated above. The plates were sealed, mixed,and incubated on an electromagnetic shaker for 10 minutes at roomtemperature, followed by 2 hours incubation at 37° C.

CFB mRNA levels were quantified by performing RT-qPCR analysis. 2 μl ofcDNA were added to a master mix containing 0.5 μl of human or cyno GAPDHTaqMan Probe, 0.5 μl human or cyno CFB probe (Hs00156060_m1) and 5 μlLightcycler 480 probe master mix (Roche Cat #04887301001) per well in a384 well plates. Real time PCR was done in a LightCycler480 Real TimePCR system (Roche). To calculate relative fold change, real-time datawere analyzed using the Delta-Delta Threshold Cycle (RelativeQuantification) (ΔΔC_(q)[RQ]) method [Schmittgen and Livak 2008] andwere normalized to control assays performed using cells transfected withPBS. For all samples, CFB mRNA levels were first normalized to GAPDH asa reference gene. Data are expressed as percent of CFB mRNA remainingrelative to average PBS control and error is expressed as standarddeviation (SD), derived from the 4 transfection replicates.

The results of single dose transfection screens and free uptake screensin PHH and PCH cells are shown in Tables 5 and 6, respectively.

TABLE 5 In Vitro Single Dose Tranfection and Free Update Screens inPrimary Human Hepatocytes Primary Human Hepatocytes Transfection PrimaryHuman Hepatocytes Free Uptake 10 nM 10 nM 1 nM 1 nM 0.1 nM 0.1 nM 0.01nM 0.01 nM 200 nM 200 nM 100 nM 100 nM 10 nM 10 nM 1 nM 1 nM Duplex NameAvg Stdev Avg Stdev Avg Stdev Avg Stdev Avg Stdev Avg Stdev Avg StdevAvg StdeV AD-1724704.1 8 2 7 3 15 3 19 2 9 3 8 2 33 5 50 8 AD-1729137.18 2 12 3 20 5 25 3 32 2 30 3 53 1 87 11 AD-1725059.1 6 2 8 1 13 4 18 410 1 10 1 27 4 61 17 AD-1727392.1 9 2 12 4 20 4 27 5 27 4 62 18 54 4 323 AD-1725408.1 7 2 11 2 16 3 18 3 16 2 15 2 46 5 68 5 AD-1727703.1 9 115 1 21 4 23 2 31 3 33 6 63 4 74 7 AD-1725478.1 10 0 13 1 20 4 27 4 20 291 6 51 7 38 5 AD-1725759.1 13 2 19 3 25 3 33 5 30 5 29 6 69 7 86 3AD-1725771.1 8 1 15 4 19 2 31 4 20 2 24 4 56 5 88 6 AD-1726051.1 16 2 163 21 4 18 2 17 5 24 2 59 7 79 9 AD-1726057.1 14 5 13 3 12 1 8 5 14 3 143 42 8 50 2 AD-1724693.1 8 2 8 2 22 0 29 3 22 1 22 2 43 2 39 16AD-1724706.1 7 1 15 3 18 2 33 5 12 2 15 1 35 2 63 9 AD-1727288.1 8 4 1 224 5 24 5 16 4 20 2 46 2 69 7 AD-1725060.1 12 1 18 5 38 4 50 5 44 3 42 273 6 96 16 AD-1725192.1 15 2 20 2 16 10 44 6 38 2 41 5 50 10 78 10AD-1725430.1 7 1 8 1 16 4 18 2 19 3 17 5 46 7 65 16 AD-1725469.1 15 3 174 32 6 41 9 43 8 41 4 77 9 96 13 AD-1725535.1 8 1 13 3 22 2 21 7 18 3 212 51 5 75 13 AD-1727981.1 13 1 16 4 21 3 28 4 27 2 32 5 61 4 89 15AD-1725772.1 10 1 16 3 22 6 31 8 20 4 25 2 54 19 107 31 AD-1728273.1 163 19 2 33 3 29 5 19 5 23 1 58 6 81 13 AD-1726058.1 7 2 15 2 15 3 18 6 135 16 2 50 7 61 21 AD-1728784.1 7 2 7 2 19 2 21 3 16 2 16 7 39 8 38 23AD-1724708.1 9 2 12 2 24 5 26 4 12 4 15 1 43 9 69 6 AD-1725193.1 12 1 181 24 6 28 6 21 2 19 4 46 6 75 10 AD-1725460.1 14 4 19 4 31 5 40 4 70 961 3 94 6 95 11 AD-1729555.1 10 2 14 3 33 7 31 8 27 8 26 1 68 7 99 28AD-1725643.1 10 1 19 5 32 9 23 7 26 3 26 3 59 2 83 5 AD-1726052.1 14 217 5 26 6 28 5 15 2 20 1 51 3 85 3 AD-1726062.1 4 1 17 1 23 11 26 8 15 518 3 45 9 48 15 AD-1726936.1 5 2 7 3 19 4 28 2 15 5 13 7 30 13 35 21AD-1724715.1 11 3 18 5 26 6 46 11 22 4 25 3 45 11 66 11 AD-1725055.1 5 28 2 20 2 21 5 17 2 20 1 47 2 74 6 AD-1725061.1 14 2 18 4 35 7 34 4 33 130 2 73 6 85 9 AD-1725194.1 13 3 14 2 26 6 23 5 16 2 16 2 40 6 62 11AD-1725462.1 13 2 14 3 34 7 36 6 46 4 29 3 69 13 80 20 AD-1725472.1 10 317 1 28 6 27 3 32 5 102 7 60 5 103 16 AD-1725644.1 13 2 21 6 31 9 31 238 4 35 8 68 13 89 18 AD-1725761.1 10 1 17 3 31 5 27 5 29 3 37 3 81 10112 8 AD-1725773.1 12 1 19 3 27 4 38 7 31 7 34 5 75 11 110 25AD-1726053.1 14 2 19 2 22 2 23 4 14 4 17 0 45 4 73 12 AD-1728302.1 10 214 3 25 8 25 8 12 3 19 4 47 9 76 17 AD-1728786.1 7 2 9 1 16 3 19 4 16 119 4 38 17 47 4 AD-1724718.1 11 1 19 4 27 4 30 5 14 3 22 2 52 2 71 5AD-1729141.1 15 4 17 2 30 5 28 5 31 7 34 4 57 5 107 19 AD-1725075.1 20 519 3 25 4 34 3 25 4 26 5 60 10 86 14 AD-1727432.1 14 2 17 2 23 3 31 5 245 104 10 56 6 82 12 AD-1729548.1 14 2 20 4 31 2 36 8 43 7 42 5 80 9 1009 AD-1725476.1 11 2 19 2 32 8 28 8 24 4 35 1 83 7 111 8 AD-1729841.1 131 21 4 29 6 25 6 36 8 41 3 74 2 111 16 AD-1725763.1 8 1 16 4 31 8 37 520 4 25 2 60 2 97 10 AD-1725777.1 14 4 18 4 18 1 21 6 24 7 27 5 65 8 9022 AD-1728276.1 15 1 23 2 34 16 29 8 18 5 22 3 44 11 94 25 AD-1730167.110 2 14 2 16 8 20 5 7 4 15 4 35 8 50 15 AD-1726937.1 6 1 9 2 8 2 20 3 112 11 1 33 2 44 9 AD-1724725.1 9 1 14 2 22 1 25 4 15 3 19 1 41 5 59 12AD-1727292.1 11 1 14 2 24 4 31 6 21 3 25 1 49 7 79 13 AD-1725095.1 12 213 1 21 4 27 5 15 2 15 1 39 4 66 6 AD-1725244.1 18 2 21 3 35 9 37 9 42 439 5 80 14 97 11 AD-1725463.1 8 2 13 3 22 8 26 9 21 3 23 3 57 9 80 10AD-1729562.1 11 2 14 2 26 5 19 4 18 3 16 3 43 3 71 7 AD-1725756.1 10 212 2 23 5 24 2 24 2 26 3 58 5 87 10 AD-1725764.1 7 1 13 2 29 7 25 4 24 5117 4 59 4 95 18 AD-1730001.1 16 2 25 11 22 5 32 7 29 5 107 15 75 9 10513 AD-1726056.1 14 2 20 2 30 9 25 3 12 2 15 2 43 7 79 17 AD-1726189.1 113 18 2 18 1 23 8 15 5 23 5 43 9 88 11 AD-1724702.1 6 1 11 2 15 4 23 7 150 15 1 36 1 50 8 AD-1724910.1 9 2 13 2 22 3 23 2 21 3 16 2 38 4 60 7AD-1727293.1 6 1 10 3 14 1 19 4 20 5 23 3 48 6 77 8 AD-1725096.1 11 3 162 30 6 34 5 22 3 25 2 55 11 81 14 AD-1729487.1 13 1 20 2 40 9 31 3 31 733 12 70 11 96 21 AD-1725464.1 9 3 14 4 26 7 27 5 21 3 28 1 64 8 91 12AD-1727713.1 11 1 16 3 32 4 34 5 26 5 28 2 67 6 97 7 AD-1725757.1 10 114 2 26 3 28 7 21 1 22 3 51 1 93 5 AD-1727986.1 7 2 12 3 29 8 27 7 17 624 2 60 10 80 13 AD-1725916.1 8 1 12 4 22 7 24 3 16 4 21 3 55 7 85 22AD-1728278.1 13 2 20 4 28 12 25 6 17 2 23 4 49 4 81 17 AD-1726207.1 9 119 5 28 6 28 3 19 5 27 6 49 9 59 7 AD-1726939.1 5 1 7 1 12 4 18 5 11 111 2 28 4 33 5 AD-1725044.1 4 1 8 2 16 4 22 7 19 3 20 2 43 9 46 10AD-1725057.1 6 3 11 2 20 2 23 6 16 2 20 2 47 9 64 14 AD-1725125.1 13 220 2 37 11 40 7 49 4 63 12 85 13 90 12 AD-1725405.1 6 2 9 2 20 6 25 8 173 21 1 57 12 66 9 AD-1725477.1 9 2 13 2 28 4 27 5 18 3 25 2 56 7 73 6AD-1727980.1 11 2 15 2 24 3 25 4 18 3 25 3 55 1 79 17 AD-1725767.1 9 217 4 41 11 37 9 24 6 35 4 66 5 87 9 AD-1730068.1 18 8 18 5 11 6 26 5 3010 43 2 77 5 92 29 AD-1730477.1 9 1 14 5 27 7 26 7 21 3 24 2 52 3 108 34

TABLE 6 In Vitro Single Dose Tranfection and Free Update Screens inPrimary Cynomolgus Hepatocytes Primary Cyno Hepatocytes TransfectionPrimary Cyno Hepatocytes Free Uptake 10 nM 10 nM 1 nM 1 nM 0.1 nM 0.1 nM0.01 nM 0.01 nM 200 nM 200 nM 100 nM 100 nM 10 nM 10 nM 1 nM 1 nM DuplexName Avg Stdev Avg Stdev Avg Stdev Avg Stdev Avg Stdev Avg Stdev AvgStdev Avg StdeV AD-1724704.1 2 1 6 1 11 2 12 3 16 1 20 2 41 2 62 5AD-1729137.1 5 0 11 1 19 1 21 1 58 1 59 6 72 2 91 6 AD-1725059.1 2 0 4 09 1 12 1 18 1 21 3 41 3 62 4 AD-1727392.1 12 1 25 2 42 1 53 3 56 2 56 480 4 93 4 AD-1725408.1 2 0 7 1 11 1 15 1 26 2 28 4 47 3 68 3AD-1727703.1 5 1 11 0 18 1 24 2 50 7 49 4 76 2 89 1 AD-1725478.1 3 0 7 112 1 17 1 28 2 33 4 56 2 72 5 AD-1725759.1 9 5 12 1 19 1 24 1 43 1 44 166 4 83 4 AD-1725771.1 4 1 10 2 17 1 23 1 31 2 33 1 56 1 75 2AD-1726051.1 6 1 11 1 16 1 23 2 41 3 44 3 70 4 87 2 AD-1726057.1 4 1 122 15 1 21 1 33 1 43 2 72 4 91 2 AD-1724693.1 5 1 11 1 22 2 30 3 36 3 372 59 3 79 4 AD-1724706.1 2 0 6 1 11 1 16 2 15 0 20 1 38 1 57 2AD-1727288.1 3 1 7 1 13 1 17 1 34 2 35 1 48 9 70 4 AD-1725060.1 6 1 16 129 3 37 2 46 1 48 3 69 4 81 1 AD-1725192.1 4 1 11 1 19 2 24 2 48 3 49 470 8 79 3 AD-1725430.1 3 1 5 1 8 1 12 1 21 1 21 1 39 2 60 3 AD-1725469.15 1 13 2 21 2 28 3 45 2 45 2 67 4 83 6 AD-1725535.1 2 0 6 1 9 1 12 1 212 23 0 45 2 67 4 AD-1727981.1 4 1 8 1 15 2 19 1 33 2 36 3 57 2 73 7AD-1725772.1 4 1 9 2 14 2 22 3 34 4 32 2 52 2 74 4 AD-1728273.1 6 1 11 117 2 22 2 42 10 39 3 63 4 81 6 AD-1726058.1 4 1 7 1 11 1 16 1 29 3 32 254 2 73 2 AD-1728784.1 4 0 9 1 19 2 24 5 31 11 38 2 61 2 79 3AD-1724708.1 3 0 8 1 13 1 17 1 20 1 21 0 44 1 62 1 AD-1725193.1 3 0 6 012 1 19 2 31 1 32 2 53 6 72 2 AD-1725460.1 5 0 12 0 22 3 29 2 68 2 63 474 5 83 3 AD-1729555.1 3 0 9 1 15 1 20 2 32 1 33 1 56 4 74 1AD-1725643.1 5 1 11 1 17 2 21 1 24 1 26 0 48 2 71 5 AD-1726052.1 5 1 101 16 2 19 1 32 1 37 2 60 4 78 3 AD-1726062.1 4 1 9 1 15 1 20 1 33 1 37 160 2 81 3 AD-1726936.1 3 0 10 1 16 0 22 1 34 3 30 3 52 3 73 3AD-1724715.1 3 0 10 1 19 2 26 0 26 1 27 2 50 2 68 1 AD-1725055.1 3 1 6 111 1 15 1 29 1 30 0 50 3 66 4 AD-1725061.1 4 0 9 1 17 1 25 1 31 2 32 251 2 69 5 AD-1725194.1 2 0 5 0 8 1 13 2 21 1 22 2 41 3 59 2 AD-1725462.14 1 10 1 18 2 24 1 36 2 36 2 55 3 74 4 AD-1725472.1 3 1 8 1 14 1 18 2 331 33 1 49 3 66 2 AD-1725644.1 6 1 13 1 21 1 24 2 40 2 40 1 60 3 78 4AD-1725761.1 4 1 8 1 15 3 19 1 33 2 33 2 55 2 70 2 AD-1725773.1 6 2 12 122 2 25 3 36 3 37 3 58 2 77 4 AD-1726053.1 4 1 7 1 12 1 13 1 25 1 29 348 4 68 3 AD-1728302.1 4 1 9 1 15 2 18 2 29 2 34 2 59 3 78 3AD-1728786.1 4 1 11 2 16 0 21 1 38 1 38 2 61 4 81 3 AD-1724718.1 3 1 111 18 1 24 2 26 1 31 3 55 4 73 2 AD-1729141.1 6 0 12 2 22 3 27 1 44 2 483 69 4 82 5 AD-1725075.1 3 0 9 1 14 1 19 2 27 1 30 3 53 5 70 2AD-1727432.1 3 0 7 0 14 2 18 1 27 2 29 2 49 4 78 10 AD-1729548.1 4 1 111 21 1 29 2 36 2 37 2 61 4 75 5 AD-1725476.1 4 1 9 1 13 2 18 0 33 1 34 057 3 70 2 AD-1729841.1 4 1 10 1 15 2 19 1 40 3 40 2 62 4 74 6AD-1725763.1 2 1 7 0 12 2 14 2 16 1 18 1 33 2 57 3 AD-1725777.1 4 1 9 117 2 21 2 29 3 35 1 58 2 71 3 AD-1728276.1 4 0 8 1 13 1 16 0 24 1 27 049 2 70 8 AD-1730167.1 3 1 8 1 13 1 17 1 25 1 29 2 50 1 75 6AD-1726937.1 2 1 7 1 11 2 17 1 21 1 24 1 47 3 73 4 AD-1724725.1 3 0 8 115 2 21 2 23 1 28 1 51 1 73 3 AD-1727292.1 3 0 6 1 12 2 16 1 29 0 31 249 2 69 3 AD-1725095.1 2 0 5 0 9 1 13 0 17 1 19 1 39 2 60 4 AD-1725244.14 1 10 1 17 1 21 2 36 3 37 2 59 3 78 3 AD-1725463.1 3 1 6 2 11 2 14 2 221 23 1 43 2 64 3 AD-1729562.1 2 0 5 1 8 1 11 1 25 1 26 1 41 3 63 5AD-1725756.1 3 1 6 1 11 2 14 2 26 3 27 1 47 2 65 5 AD-1725764.1 2 0 6 112 2 16 2 25 1 26 2 44 3 64 5 AD-1730001.1 5 1 12 1 22 2 24 3 30 1 35 258 3 73 4 AD-1726056.1 4 1 8 1 13 2 14 1 24 1 28 2 50 4 70 3AD-1726189.1 7 1 18 3 24 1 30 2 53 3 58 3 79 4 94 6 AD-1724702.1 4 0 101 16 2 22 2 31 2 33 2 57 1 77 4 AD-1724910.1 3 1 8 1 12 1 15 1 32 1 31 152 1 70 4 AD-1727293.1 4 1 8 0 16 2 22 3 40 1 43 2 66 5 80 3AD-1725096.1 3 1 7 1 14 1 20 2 22 1 24 1 45 2 71 3 AD-1729487.1 7 1 13 124 4 33 6 47 1 49 3 70 4 85 4 AD-1725464.1 2 0 6 1 13 2 19 2 23 1 25 244 4 64 2 AD-1727713.1 3 1 7 1 13 1 18 1 30 1 33 2 57 1 77 2AD-1725757.1 3 0 6 1 10 2 14 1 24 2 24 1 46 1 65 3 AD-1727986.1 3 1 6 114 1 17 2 24 1 27 2 47 4 65 4 AD-1725916.1 3 1 7 1 12 1 14 0 21 1 24 147 4 67 6 AD-1728278.1 3 0 8 1 12 2 14 1 25 2 30 2 54 2 72 5AD-1726207.1 4 1 10 1 19 3 25 2 25 1 30 2 53 3 76 5 AD-1726939.1 3 1 6 012 2 17 0 21 1 23 1 46 4 74 5 AD-1725044.1 3 0 7 2 14 1 16 5 36 2 32 150 2 75 3 AD-1725057.1 3 0 7 1 14 2 20 1 42 3 38 1 60 4 80 3AD-1725125.1 8 1 23 3 39 5 53 5 61 2 62 2 82 3 98 4 AD-1725405.1 2 1 7 113 1 19 2 22 2 23 1 43 1 71 4 AD-1725477.1 3 0 7 1 11 4 18 2 32 3 34 156 2 77 4 AD-1727980.1 4 1 9 0 16 1 20 3 29 2 31 3 51 3 74 7AD-1725767.1 6 1 12 1 23 2 27 1 41 1 42 2 64 7 87 4 AD-1730068.1 5 1 110 18 2 19 2 35 2 36 2 63 5 84 6 AD-1730477.1 3 0 10 2 13 2 18 1 32 3 331 51 3 77 7

Example 4. In Vivo Screening of dsRNA Duplexes

Single Dose Study (1 mg/kg)

Duplexes of interest, identified from the above in vitro studies, wereevaluated in vivo. In particular, at pre-dose day −21 wild-type mice(C57BL/6) were transduced with 2×10¹¹ viral particles of anadeno-associated virus 8 (AAV8) vector encoding human CFB intravenouslyvia retro-orbital delivery. In particular, mice were administered anAAV8 encoding a portion of human CFB mRNA encoding the open readingframe and 3′ UTR of human CFB mRNA referenced as NM_001710.5, referredto as VCAV-07851.AAV8.HsCFB.

At day 0, groups of three mice were subcutaneously administered a single1 mg/kg dose of the duplexes of interest or phosphate-buffered saline(PBS). Table 7 provides the treatment design and provides the duplexesof interest. On day 0 predose and day 7 post-dose, K₂EDTA plasma werecollected and samples were analyzed for human CFB protein by ELISA(Assay Pro #EF7001). All plasma samples were diluted 1:1000 in 1×diluent and ELISA was conducted according to manufacturer's protocol.Data was interpolated using a 4-parameter logistic curve using GraphPadPrism software.

For all samples, human CFB protein levels were normalized to individualanimals' day 0 level to calculate relative % human CFB proteinremaining. For each group, the mean % CFB protein remaining wascalculated (±standard deviation [SD]).

The data were expressed as percent of baseline value and presented asmean plus standard deviation. The results, listed in Table 8 and shownin FIG. 2 , demonstrate that the exemplary duplex agents testedeffectively reduce the level of the human CFB protein in vivo.

TABLE 7 Treatment Groups # of Dose Target Group Treatment animals (mpk)region  1 PBS 3 NA  2 AD-1726057.3 3 1 2242-2264  3 AD-1725763.3 3 11828-1850  4 AD-1725777.3 3 1 1842-1864  5 AD-1725057.3 3 1  995-1017  6AD-1725096.3 3 1 1034-1056  7 AD-1728786.3 3 1  641-663  8 AD-1725059.33 1  997-1019  9 AD-1728276.3 3 1 2391-2413 10 AD-1728278.3 3 12393-2415 11 AD-1726936.3 3 1  640-662 12 AD-1725472.3 3 1 1473-1495 13AD-1724715.3 3 1  504-526 14 AD-1727292.3 3 1 1145-1167 15 AD-1730477.33 1 2453-2475 17 AD-1727288.3 3 1 1141-1163 18 AD-1730167.3 3 12438-2460 19 AD-1725408.3 3 1 1389-1411 20 AD-1725761.3 3 1 1826-1848

TABLE 8 In Vivo Screen % CFB remaining Group Group Treatment (Individualanimals) Mean STDEV  1 PBS 100.01 106.18 98.65 101.61 4.01  2AD-1726057.3 67.11 72.07 68.00 69.06 2.64  3 AD-1725763.3 35.75 44.8231.92 37.50 6.63  4 AD-1725777.3 80.87 67.57 72.06 73.50 6.77  5AD-1725057.3 60.58 72.93 70.28 67.93 6.50  6 AD-1725096.3 68.25 57.3465.02 63.54 5.61  7 AD-1728786.3 68.9 65.6 69.2 67.91 1.99  8AD-1725059.3 62.9 62.2 64.2 63.10 0.99  9 AD-1728276.3 49.2 26.3 38.738.06 11.43 10 AD-1728278.3 60.5 56.1 58.1 58.24 2.16 11 AD-1726936.338.4 62.2 50.0 50.21 11.92 12 AD-1725472.3 47.5 76.4 64.5 62.78 14.53 13AD-1724715.3 51.69 47.15 66.83 55.23 10.30 14 AD-1727292.3 43.51 50.5353.90 49.31 5.30 15 AD-1730477.3 50.08 50.96 ND 50.52 0.62 17AD-1727288.3 67.03 56.42 42.49 55.31 12.31 18 AD-1730167.3 54.47 56.2452.30 54.34 1.97 19 AD-1725408.3 56.03 39.87 58.80 51.56 10.22 20AD-1725761.3 71.73 75.44 52.36 66.51 12.39

Equivalents

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments and methods described herein. Such equivalents are intendedto be encompassed by the scope of the following claims.

Informal Sequence Listing

>gi|1732746151|ref|NM_001710.6 Homo sapienscomplement factor B (CFB), mRNA SEQ ID NO: 1GGGAAGGGAATGTGACCAGGTCTAGGTCTGGAGTTTCAGCTTGGACACTGAGCCAAGCAGACAAGCAAAGCAAGCCAGGACACACCATCCTGCCCCAGGCCCAGCTTCTCTCCTGCCTTCCAACGCCATGGGGAGCAATCTCAGCCCCCAACTCTGCCTGATGCCCTTTATCTTGGGCCTCTTGTCTGGAGGTGTGACCACCACTCCATGGTCTTTGGCCCGGCCCCAGGGATCCTGCTCTCTGGAGGGGGTAGAGATCAAAGGCGGCTCCTTCCGACTTCTCCAAGAGGGCCAGGCACTGGAGTACGTGTGTCCTTCTGGCTTCTACCCGTACCCTGTGCAGACACGTACCTGCAGATCTACGGGGTCCTGGAGCACCCTGAAGACTCAAGACCAAAAGACTGTCAGGAAGGCAGAGTGCAGAGCAATCCACTGTCCAAGACCACACGACTTCGAGAACGGGGAATACTGGCCCCGGTCTCCCTACTACAATGTGAGTGATGAGATCTCTTTCCACTGCTATGACGGTTACACTCTCCGGGGCTCTGCCAATCGCACCTGCCAAGTGAATGGCCGATGGAGTGGGCAGACAGCGATCTGTGACAACGGAGCGGGGTACTGCTCCAACCCGGGCATCCCCATTGGCACAAGGAAGGTGGGCAGCCAGTACCGCCTTGAAGACAGCGTCACCTACCACTGCAGCCGGGGGCTTACCCTGCGTGGCTCCCAGCGGCGAACGTGTCAGGAAGGTGGCTCTTGGAGCGGGACGGAGCCTTCCTGCCAAGACTCCTTCATGTACGACACCCCTCAAGAGGTGGCCGAAGCTTTCCTGTCTTCCCTGACAGAGACCATAGAAGGAGTCGATGCTGAGGATGGGCACGGCCCAGGGGAACAACAGAAGCGGAAGATCGTCCTGGACCCTTCAGGCTCCATGAACATCTACCTGGTGCTAGATGGATCAGACAGCATTGGGGCCAGCAACTTCACAGGAGCCAAAAAGTGTCTAGTCAACTTAATTGAGAAGGTGGCAAGTTATGGTGTGAAGCCAAGATATGGTCTAGTGACATATGCCACATACCCCAAAATTTGGGTCAAAGTGTCTGAAGCAGACAGCAGTAATGCAGACTGGGTCACGAAGCAGCTCAATGAAATCAATTATGAAGACCACAAGTTGAAGTCAGGGACTAACACCAAGAAGGCCCTCCAGGCAGTGTACAGCATGATGAGCTGGCCAGATGACGTCCCTCCTGAAGGCTGGAACCGCACCCGCCATGTCATCATCCTCATGACTGATGGATTGCACAACATGGGCGGGGACCCAATTACTGTCATTGATGAGATCCGGGACTTGCTATACATTGGCAAGGATCGCAAAAACCCAAGGGAGGATTATCTGGATGTCTATGTGTTTGGGGTCGGGCCTTTGGTGAACCAAGTGAACATCAATGCTTTGGCTTCCAAGAAAGACAATGAGCAACATGTGTTCAAAGTCAAGGATATGGAAAACCTGGAAGATGTTTTCTACCAAATGATCGATGAAAGCCAGTCTCTGAGTCTCTGTGGCATGGTTTGGGAACACAGGAAGGGTACCGATTACCACAAGCAACCATGGCAGGCCAAGATCTCAGTCATTCGCCCTTCAAAGGGACACGAGAGCTGTATGGGGGCTGTGGTGTCTGAGTACTTTGTGCTGACAGCAGCACATTGTTTCACTGTGGATGACAAGGAACACTCAATCAAGGTCAGCGTAGGAGGGGAGAAGCGGGACCTGGAGATAGAAGTAGTCCTATTTCACCCCAACTACAACATTAATGGGAAAAAAGAAGCAGGAATTCCTGAATTTTATGACTATGACGTTGCCCTGATCAAGCTCAAGAATAAGCTGAAATATGGCCAGACTATCAGGCCCATTTGTCTCCCCTGCACCGAGGGAACAACTCGAGCTTTGAGGCTTCCTCCAACTACCACTTGCCAGCAACAAAAGGAAGAGCTGCTCCCTGCACAGGATATCAAAGCTCTGTTTGTGTCTGAGGAGGAGAAAAAGCTGACTCGGAAGGAGGTCTACATCAAGAATGGGGATAAGAAAGGCAGCTGTGAGAGAGATGCTCAATATGCCCCAGGCTATGACAAAGTCAAGGACATCTCAGAGGTGGTCACCCCTCGGTTCCTTTGTACTGGAGGAGTGAGTCCCTATGCTGACCCCAATACTTGCAGAGGTGATTCTGGCGGCCCCTTGATAGTTCACAAGAGAAGTCGTTTCATTCAAGTTGGTGTAATCAGCTGGGGAGTAGTGGATGTCTGCAAAAACCAGAAGCGGCAAAAGCAGGTACCTGCTCACGCCCGAGACTTTCACATCAACCTCTTTCAAGTGCTGCCCTGGCTGAAGGAGAAACTCCAAGATGAGGATTTGGGTTTTCTATAAGGGGTTTCCTGCTGGACAGGGGCGTGGGATTGAATTAAAACAGCTGCGACAACA>gi|218156288|ref|NM_008198.2|Mus musculus complementfactor B (Cfb), transcript variant 1, mRNA SEQ ID NO: 2GCTCCATCACACAGTCCATGGAAAGACTGATCTTTTAAATTGGGGGTAGTGGAGGTGGTGGTCTGTGCTTGTTAGGAGGGGTCTGGGGGCTAAGAGGGAGCTTTGAAAGGGAAGTTCTGGCCCTTGGTCAGTCAAGGGTGGGGCTCACATAGTTTCTGTTTCCTCAGTTGGCAGTTCAGCTGGGGCCCTCCTTCATGAATGTTCCGGGAAGCAGTGGCTGCGTGCGCAGGGTAGGCTGGCCAGGCTGCAGATGCCAGAGCAGATTGCATAAAAGGTTAGGGGACAGTGGGAAAGGGGTGTAGCCAGATCCAGCATTTGGGTTTCAGTTTGGACAGGAGGTCAAATAGGCACCCAGAGTGACCTGGAGAGGGCTTTGGGCCACTGGACTCTCTGGTGCTTTCCATGACAATGGAGAGCCCCCAGCTCTGCCTCGTCCTCTTGGTCTTAGGCTTCTCCTCTGGAGGTGTGAGCGCAACTCCAGTGCTTGAGGCCCGGCCCCAAGTCTCCTGCTCTCTGGAGGGAGTAGAGATCAAAGGCGGCTCCTTTCAACTTCTCCAAGGCGGTCAGGCCCTGGAGTACCTATGTCCCTCTGGCTTCTACCCATACCCCGTGCAGACTCGAACCTGCAGATCCACAGGCTCCTGGAGCGACCTGCAGACCCGAGACCAAAAGATTGTCCAGAAGGCGGAATGCAGAGCAATACGCTGCCCACGACCGCAGGACTTTGAAAATGGGGAATTCTGGCCCCGGTCCCCCTTCTACAACCTGAGTGACCAGATTTCTTTTCAATGCTATGATGGTTACGTTCTCCGGGGCTCTGCTAATCGCACCTGCCAAGAGAATGGCCGGTGGGATGGGCAAACAGCAATTTGTGATGATGGAGCTGGATACTGTCCCAATCCCGGTATTCCTATTGGGACAAGGAAGGTGGGTAGCCAATACCGCCTTGAAGACATTGTTACTTACCACTGCAGCCGGGGACTTGTCCTGCGTGGCTCCCAGAAGCGAAAGTGTCAAGAAGGTGGCTCATGGAGTGGGACAGAGCCTTCCTGCCAAGATTCCTTCATGTATGACAGCCCTCAAGAAGTGGCCGAAGCATTCCTATCCTCCCTGACAGAGACCATCGAAGGAGCCGATGCTGAGGATGGGCACAGCCCAGGAGAACAGCAGAAGAGGAAGATTGTCCTAGACCCCTCGGGCTCCATGAATATCTACCTGGTGCTAGATGGATCAGACAGCATCGGAAGCAGCAACTTCACAGGGGCTAAGCGGTGCCTCACCAACTTGATTGAGAAGGTGGCGAGTTACGGGGTGAGGCCACGATATGGTCTCCTGACATATGCTACAGTCCCCAAAGTGTTGGTCAGAGTGTCTGATGAGAGGAGTAGCGATGCCGACTGGGTCACAGAGAAGCTCAACCAAATCAGTTATGAAGACCACAAGCTGAAGTCAGGGACCAACACCAAGAGGGCTCTCCAGGCTGTGTATAGCATGATGAGCTGGGCAGGGGATGCCCCGCCTGAAGGCTGGAATAGAACCCGCCATGTCATCATCATTATGACTGATGGCTTGCACAACATGGGTGGAAACCCTGTCACTGTCATTCAGGACATCCGAGCCTTGCTGGACATCGGCAGGGATCCCAAAAATCCCAGGGAGGATTACCTGGATGTGTATGTGTTTGGGGTCGGGCCTCTGGTGGACTCCGTGAACATCAATGCCTTAGCTTCCAAAAAGGACAATGAGCATCATGTGTTTAAAGTCAAGGATATGGAAGACCTGGAGAATGTTTTCTACCAAATGATTGATGAAACCAAATCTCTGAGTCTCTGTGGCATGGTGTGGGAGCATAAAAAAGGCAACGATTATCATAAGCAACCATGGCAAGCCAAGATCTCAGTCACTCGCCCTCTGAAAGGACATGAGACCTGTATGGGGGCCGTGGTGTCTGAGTACTTCGTGCTGACAGCAGCGCACTGCTTCATGGTGGATGATCAGAAACATTCCATCAAGGTCAGCGTGGGGGGTCAGAGGCGGGACCTGGAGATTGAAGAGGTCCTGTTCCACCCCAAATACAATATTAATGGGAAAAAGGCAGAAGGGATCCCTGAGTTCTATGATTATGATGTGGCCCTAGTCAAGCTCAAGAACAAGCTCAAGTATGGCCAGACTCTCAGGCCCATCTGTCTCCCCTGCACGGAGGGAACCACACGAGCCTTGAGGCTTCCTCAGACAGCCACCTGCAAGCAGCACAAGGAACAGTTGCTCCCTGTGAAGGATGTCAAAGCTCTGTTTGTATCTGAGCAAGGGAAGAGCCTGACTCGGAAGGAGGTGTACATCAAGAATGGGGACAAGAAAGCCAGTTGTGAGAGAGATGCTACAAAGGCCCAAGGCTATGAGAAGGTCAAAGATGCCTCTGAGGTGGTCACTCCACGGTTCCTCTGCACAGGAGGGGTGGATCCCTATGCTGACCCCAACACATGCAAAGGAGATTCCGGGGGCCCTCTCATTGTTCACAAGAGAAGCCGCTTCATTCAAGTTGGTGTGATTAGCTGGGGAGTAGTAGATGTCTGCAGAGACCAGAGGCGGCAACAGCTGGTACCCTCTTATGCCCGGGACTTCCACATCAACCTCTTCCAGGTGCTGCCCTGGCTAAAGGACAAGCTCAAAGATGAGGATTTGGGTTTTCTATAAAGAGCTTCCTGCAGGGAGAGTGTGAGGACAGATTAAAGCAGTTACAATAACAAAAAAAAAAAAAAAAAAAAAA>gi|218156290|ref|NM_001142706.1 Mus musculus complementfactor B (Cfb), transcript variant 2, mRNA SEQ ID NO: 3GCTCCATCACACAGTCCATGGAAAGACTGATCTTTTAAATTGGGGGTAGTGGAGGTGGTGGTCTGTGCTTGTTAGGAGGGGTCTGGGGGCTAAGAGGGAGCTTTGAAAGGGAAGTTCTGGCCCTTGGTCAGTCAAGGGTGGGGCTCACATAGTTTCTGTTTCCTCAGTTGGCAGTTCAGCTGGGGCCCTCCTTCATGAATGTTCCGGGAAGCAGTGGCTGCGTGCGCAGGGTAGGCTGGCCAGGCTGCAGATGCCAGAGCAGATTGCATAAAAGGTTAGGGGACAGTGGGAAAGGGGTGTAGCCAGATCCAGCATTTGGGTTTCAGTTTGGACAGGAGGTCAAATAGGCACCCAGAGTGACCTGGAGAGGGCTTTGGGCCACTGGACTCTCTGGTGCTTTCCATGACAATGGAGAGCCCCCAGCTCTGCCTCGTCCTCTTGGTCTTAGGCTTCTCCTCTGGAGGTGTGAGCGCAACTCCAGTGCTTGAGGCCCGGCCCCAAGTCTCCTGCTCTCTGGAGGGAGTAGAGATCAAAGGCGGCTCCTTTCAACTTCTCCAAGGCGGTCAGGCCCTGGAGTACCTATGTCCCTCTGGCTTCTACCCATACCCCGTGCAGACTCGAACCTGCAGATCCACAGGCTCCTGGAGCGACCTGCAGACCCGAGACCAAAAGATTGTCCAGAAGGCGGAATGCAGAGCAATACGCTGCCCACGACCGCAGGACTTTGAAAATGGGGAATTCTGGCCCCGGTCCCCCTTCTACAACCTGAGTGACCAGATTTCTTTTCAATGCTATGATGGTTACGTTCTCCGGGGCTCTGCTAATCGCACCTGCCAAGAGAATGGCCGGTGGGATGGGCAAACAGCAATTTGTGATGATGGAGCTGGATACTGTCCCAATCCCGGTATTCCTATTGGGACAAGGAAGGTGGGTAGCCAATACCGCCTTGAAGACATTGTTACTTACCACTGCAGCCGGGGACTTGTCCTGCGTGGCTCCCAGAAGCGAAAGTGTCAAGAAGGTGGCTCATGGAGTGGGACAGAGCCTTCCTGCCAAGATTCCTTCATGTATGACAGCCCTCAAGAAGTGGCCGAAGCATTCCTATCCTCCCTGACAGAGACCATCGAAGGAGCCGATGCTGAGGATGGGCACAGCCCAGGAGAACAGCAGAAGAGGAAGATTGTCCTAGACCCCTCGGGCTCCATGAATATCTACCTGGTGCTAGATGGATCAGACAGCATCGGAAGCAGCAACTTCACAGGGGCTAAGCGGTGCCTCACCAACTTGATTGAGAAGGTGGCGAGTTACGGGGTGAGGCCACGATATGGTCTCCTGACATATGCTACAGTCCCCAAAGTGTTGGTCAGAGTGTCTGATGAGAGGAGTAGCGATGCCGACTGGGTCACAGAGAAGCTCAACCAAATCAGTTATGAAGACCACAAGCTGAAGTCAGGGACCAACACCAAGAGGGCTCTCCAGGCTGTGTATAGCATGATGAGCTGGGCAGGGGATGCCCCGCCTGAAGGCTGGAATAGAACCCGCCATGTCATCATCATTATGACTGATGGCTTGCACAACATGGGTGGAAACCCTGTCACTGTCATTCAGGACATCCGAGCCTTGCTGGACATCGGCAGGGATCCCAAAAATCCCAGGGAGGATTACCTGGATGTGTATGTGTTTGGGGTCGGGCCTCTGGTGGACTCCGTGAACATCAATGCCTTAGCTTCCAAAAAGGACAATGAGCATCATGTGTTTAAAGTCAAGGATATGGAAGACCTGGAGAATGTTTTCTACCAAATGATTGATGAAACCAAATCTCTGAGTCTCTGTGGCATGGTGTGGGAGCATAAAAAAGGCAACGATTATCATAAGCAACCATGGCAAGCCAAGATCTCAGTCACTCGCCCTCTGAAAGGACATGAGACCTGTATGGGGGCCGTGGTGTCTGAGTACTTCGTGCTGACAGCAGCGCACTGCTTCATGGTGGATGATCAGAAACATTCCATCAAGGTCAGCGTGGGGGGTCAGAGGCGGGACCTGGAGATTGAAGAGGTCCTGTTCCACCCCAAATACAATATTAATGGGAAAAAGGCAGAAGGGATCCCTGAGTTCTATGATTATGATGTGGCCCTAGTCAAGCTCAAGAACAAGCTCAAGTATGGCCAGACTCTCAGGCCCATCTGTCTCCCCTGCACGGAGGGAACCACACGAGCCTTGAGGCTTCCTCAGACAGCCACCTGCAAGCAGCACAAGGAACAGTTGCTCCCTGTGAAGGATGTCAAAGCTCTGTTTGTATCTGAGCAAGGGAAGAGCCTGACTCGGAAGGAGGTGTACATCAAGAATGGGGACAAGCCAGTTGTGAGAGAGATGCTACAAAGGCCCAAGGCTATGAGAAGGTCAAAGATGCCTCTGAGGTGGTCACTCCACGGTTCCTCTGCACAGGAGGGGTGGATCCCTATGCTGACCCCAACACATGCAAAGGAGATTCCGGGGGCCCTCTCATTGTTCACAAGAGAAGCCGCTTCATTCAAGTTGGTGTGATTAGCTGGGGAGTAGTAGATGTCTGCAGAGACCAGAGGCGGCAACAGCTGGTACCCTCTTATGCCCGGGACTTCCACATCAACCTCTTCCAGGTGCTGCCCTGGCTAAAGGACAAGCTCAAAGATGAGGATTTGGGTTTTCTATAAAGAGCTTCCTGCAGGGAGAGTGTGAGGACAGATTAAAGCAGTTACAATAACAAAAAAAAAAAAAAAAAAAAAA>gi|218156284|ref|NM_212466.3 Rattus norvegicus complementfactor B (Cfb), mRNA SEQ ID NO: 4CAGCAGGGGCCCTCCTTCATGAATGTTCCGGGAAGCAGCGTCTGTGCAGGGTAGGTTGGCCAGGCTGCAGGTGCCAGAGCAGATTGCATAAAAGGTTAGGGGCCGGTGGGAAAGGGGTGTAGCCAGATCCAGCACTGGAGTTTCAGTCTGGACAGCAAGTCAAGTAGCCACCCAGAGTGAACTGGAAAGGGCTTTTGGCCACGGGCTTTCCATGACAATGGAGGGTCCCCAGCTCTGCTTAGTCCTCTTGGTCTTAGGCCTCTCCTCCGGAGGTGTGAGCGCAACTCCAGTGCTTGAGGCCCGGCCCCAGGTCTCTTGCTCTCTGGAGGGAGTAGAGATCAAAGGCGGCTCCTTCCAACTTCTCCAAGACGGTCAGGCCCTGGAGTACCTGTGTCCCTCTGGCTTCTACCCATACCCTGTGCAGACTCGAACCTGCAAATCCACAGGCTCCTGGAGTGTCCTCCAGACCCGGGACCAAAAGATTGTCAAGAAGGCAGAATGCAGAGCAATACGCTGCCCACGACCACAGGACTTTGAAAATGGGGAGTTCTGGCCCCGGTCCCCCTACTACAACCTGAGTGATCAGATTTCTTTTCAATGCTATGATGGCTACACTCTCCGGGGCTCTGCTAATCGCACCTGCCAAGAGAATGGCCGGTGGGATGGGCAAACAGCAATCTGTGATGATGGAGCGGGATACTGTCCCAACCCGGGTATTCCTATTGGGACAAGGAAGGTGGGAAGCCAGTACCGTCTTGAAGACACTGTCACTTACCACTGTAGTCGGGGACTTGTCCTACGTGGCTCCCAGCAGCGAAGGTGCCAGGAAGGTGGCTCGTGGAGTGGGACAGAGCCTTCCTGCCAAGATTCCTTCATGTACGACAGCCCTCAAGAGGTGGCCGAAGCATTTCTATCCTCCCTGACAGAGACCATCGAAGGAGCAGATGCGGAGGATGGGCACAGCCCAGGGGAACAGCAGAAGAGGAAGATTATCCTGGACCCCTCGGGCTCCATGAATATCTACATGGTGCTGGATGGATCCGACAGCATCGGGGCCAGCAACTTCACAGGGGCCAAGCGGTGTCTCGCCAACTTGATTGAGAAGGTGGCGAGTTATGGGGTGAAGCCAAGATACGGTCTAGTGACATATGCCACAGTCCCCAAAGTCTTGGTCAGAGTGTCTGAGGAGAGGAGTAGTGATGCCGACTGGGTCACAGAGAAGCTCAACCAAATCAGTTATGAAGACCACAAGCTGAAGTCAGGGACCAACACCAAGAAGGCTCTCCAGGCTGTATACAGCATGATGAGCTGGCCAGGGGATGCTCCGCCTGAAGGCTGGAATCGAACCCGCCACGTCATCATCATCATGACTGATGGCTTGCACAACATGGGTGGAGACCCTGTCACTGTCATTGAGGACATCCGAGACTTGCTGGATATTGGCAGGGATCGCAAAAATCCCCGGGAGGATTATTTGGATGTGTATGTGTTTGGGGTCGGGCCTCTGGTGGACCCTGTGAACATCAATGCCTTGGCTTCCAAAAAGAACAATGAGCAGCATGTGTTCAAGGTCAAGGACATGGAGGATCTGGAGAACGTCTTCTACAAAATGATCGATGAAACCAAATCTCTGGGTCTCTGTGGCATGGTGTGGGAGCATCAGAAAGGCGGTGATTATTACAAGCAACCATGGCAAGCCAAGATCTCAGTCACTCGTCCTCTGAAAGGACATGAGAACTGTATGGGGGCCGTGGTGTCCGAGTACTTCGTGCTGACAGCAGCGCATTGCTTCACAGTGGAAGATCAGAAACACTCCATCAAGGTCAACGTGGAGGGGAAAAGGCGGGACCTGGAGATTGAAGAGGTCCTGTTCCACCCTAATTACGACATCAATGGGAAAAAGGCAGAAGGAATCTCTGAGTTCTATGACTATGATGTTGCCCTCATCAAGCTCAAGACCAAGCTGAAGTACAGCCAGACTCTCAGGCCCATCTGTCTCCCCTGCACAGAGGGAACCACCCGAGCCTTGCGGCTTCCTCAGACAGCCACCTGCAAACAGCACAAGGAAGAGTTGCTCCCTATGAAGGACGTCAAAGCTCTGTTTGTATCCGAGGAAGGGAAGAAGCTGACCCGGAAGGAGGTGTACATCAAGAATGGGGAAAAGAAAGCCAGTIGTGAGAGAGATGCTACAAAGGCCCAAGGCTATGAGAAGGTCAAAGTTGCCTCTGAGGTGGTCACCCCCAGGTTCCTGTGCACCGGAGGGGTAGATCCCTATGCTGACCCCAACACATGCAAAGGAGACTCCGGGGGCCCTCTCATTGTTCACAAGAGAAGCCGCTTCATTCAAGTTGGTGTGATCAGCTGGGGAGTAGTGGATGTCTGCAAAGACCCGAGGCGGCAACAGTTGGTGCCCTCCTATGCCCGGGACTTCCACATCAATCTCTTCCAGGTGCTGCCCTGGCTAAAGGAGAAGCTCAAAGACGAGGACTTGGGTTTCTTATAAGGAGCTTCCTGCTGGGAGGGTGAGGGCAGATTAAAGCAGCTACAATACAAATACAAAAAAAAAAAAAAAA>gi|57114201|ref|NM_001009169.1|Pan troglodytescomplement factor B (CFB), mRNA SEQ ID NO: 5CCCAGGCCCAGCTTCTCTCCTGCCTTCCAACGCCATGGGGAGCAATCTCAGCCCCCAACTCTGCCTGATGCCCTTCATCTTGGGCCTCTTGTCTGGAGGTGTGACCACCACTCCATGGCCTTTGGCCCAGCCCCAGGAATCCTGCTCTCTGGAGGGGGTAGAGATCAAAGGCGGCTCCTTCCGACTTCTCCAAGAGGGCCAGGCACTGGAGTACGTGTGTCCTTCTGGCTTCTACCCGTACCCTGTGCAGACACGTACCTGCAGATCTACGGGGTCCTGGAGCACCCTGAAGACTCAAGTCCAAAAGACTGTCAGGAAGGCAGAGTGCAGAGCAATCCACTGTCCAAGACCACACGACTTCGAGAACGGGGAATACTGGCCCCGGTCTCCCTACTACAATGTGAGTGATGAGATCTCTTTCCACTGCTATGACGGTTACACTCTCCGGGGCTCTGCCAATCGCACCTGCCAAGTGAATGGCCGGTGGAGTGGGCAGACAGCGATCTGTGACAACGGAGCGGGGTACTGCTCCAACCCGGGCATCCCCATTGGCACAAGGAAGGTGGGCAGCCAGTACCGCCTTGAAGACAGCGTCACCTACCACTGCAGCCGGGGGCTTACCCTGCGTGGCTCCCAGCGGCGAACGTGTCAGGAAGGTGGCTCTTGGAGCGGGACGGAGCCTTCTTGCCAAGACTCCTTCATGTACGACACCCCTCAAGAGGTGGCCGAAGCTTTCCTGTCTTCCCTGACAGAGACCATAGAAGGAGTCGATGCTGAGGATGGGCACGGCCCAGGGGAACAACAGAAGCGGAAGATCGTCCTGGACCCTTCAGGCTCCATGAACATCTACCTGGTGCTAGATGGATCAGACAGCATTGGGGCCAGCAACTTCACAGGAGCCAAAAAGTGTCTAGTCAACTTAATTGAGAAGGTGGCAAGTTATGGTGTGAAGCCAAGATATGGTCTAGTGACATATGCCACACACCCCAAAATTTGGGTCAAAGTGTCTGATCCAGACAGCAGTAATGCAGACTGGGTCACGAAGCAGCTCAATGAAATCAATTATGAAGACCACAAGTTGAAGTCAGGGACTAACACCAAGAAGGCCCTCCAGGCAGTGTACAGCATGATGAGCTGGCCAGATGACATCCCTCCTGAAGGCTGGAACCGCACCCGCCATGTCATCATCCTCATGACTGATGGATTGCACAACATGGGCGGGGACCCAATTACTGTCATTGATGAGATCCGGGACTTGCTATACATTGGCAAGGATCGCAAAAACCCAAGGGAGGATTATCTGGATGTCTATGTGTTTGGGGTCGGGCCTTTGGTGAACCAAGTGAACATCAATGCTTTGGCTTCCAAGAAAGACAATGAGCAACATGTGTTCAAAGTCAAGGATATGGAAAACCTGGAAGATGTTTTCTACCAAATGATTGATGAAAGCCAGTCTCTGAGTCTCTGTGGCATGGTTTGGGAACACAGGAAGGGTACCGATTACCACAAGCAACCATGGCAAGCCAAGATCTCAGTCATTCGCCCTTCAAAGGGACACGAGAGCTGTATGGGGGCTGTGGTGTCTGAGTACTTTGTGCTGACAGCAGCACACTGTTTCACTGTGGATGACAAGGAACACTCAATCAAGGTCAGCGTAGGAGGGGAGAAGCGGGACCTGGATATGACTATGACGTTGCCCTGATCAAGCTCAAGAATAAGCTGAAATATGGCCAGACTATCAGGCCCATTTGTCTCCCCTGCACCGAGGGAACAACTCGAGCTTTGAGGCTTCCTCCAACTACCACTTGCCAGCAACAAAAGGAAGAGCTGCTCCCTGCACAGGATATCAAAGCTCTGTTTGTGTCTGAGGAGGAGAAAAAGCTGACTCGGAAGGAGGTCTACATCAAGAATGGGGATAAGAAAGGCAGCTGTGAGAGAGATGCTCAATATGCCCCAGGCTATGACAAAGTCAAGGACATCTCAGAGGTGGTCACCCCTCGGTTCCTTTGTACTGGAGGAGTGAGTCCCTATGCTGACCCCAATACTTGCAGAGGTGATTCTGGCGGCCCCTTGATAGTTCACAAAAGAAGTCGTTTCATTCAAGTTGGTGTAATCAGCTGGGGAGTAGTGGATGTCTGCAAAAACCAGAAGCGGCAAAAGCAGGTACCTGCTCACGCCCGAGACTTTCACATCAACCTCTTTCAAGTGCTGCCCTGGCTGAAGGAGAAACTCCAAGATGAGGATTTGGGTTTTCTATAAGGGGTMacaca fascicularis Complement Factor B >ENSMMUP00000000985 [mRNA] locus = scaffold3881:47830:53620:- SEQ ID NO: 6ATGGGGAGCAGTCTCAGCCCCCAGCTCTACCTGATGCCCTTCATCTTGGGCCTCTTATCTGCAGGTGTGACCACCACTCCATTGTCTTCGGCCCAGCCTCAAGGATCCTGCTCTCTGGAGGGGGTAGAGATCAAAGGTGGCTCCTTCCGACTTCTCCAAGAGGGCCAGGCACTGGAATACGTGTGTCCTTCTGGCTTCTACCCGTACCCTGTGCAGACACGTACCTGCAGATCCACGGGGTCCTGGAGCACCCTGCAGACTCAAGATCGAAAAACTGTCAAGAAGGCAGAGTGCAGAGCAATCCGCTGTCCACGACCACAGGACTTCGAGAACGGGGAATACCGGCCCCGGTCTCCCTACTACAATGTGAGTGATGAGATCTCTTTCCACTGCTATGACGGTTACACTCTCCGGGGCTCTGCCAATCGCACCTGCCAAGTGAATGGCCGGTGGAGTGGGCAGACAGCGATCTGTGACAACGGAGCGGGGTACTGCTCCAACCCAGGCATCCCCATTGGCACAAGGAAGGTGGGCAGCCGGTACCGCCTTGAAGACAGCGTCACCTACCACTGCAGCCGGGGGCTTACCCTGCGTGGCTCCCAGCGGCGAACATGTCAGGAAGGTGGCTCTTGGAGCGGGACGGAGCCTTCCTGCCAAGACTCCTTCATGTACGACACCCCTCAAGAGGTGGCCGAAGCTTTCCTGTCTTCCCTGACGGAGACCATAGAAGGAGTCGATGCCGAGGATGGGCACAGCCCAGGGGAACAACAGAAGCGGAGGATCATCCTAGACCCTTCAGGCTCCATGAACATCTACCTGGTGCTAGATGGATCAGACAGCATTGGGGCCGGCAACTTCACAGGAGCCAAAAAGTGTCTAGTCAACTTAATTGAGAAGGTGGCAAGTTATGGTGTGAAGCCAAGATATGCTCTAGTGACATATGCCACATACCCCAGAATTTGGGTCAAAGTGTCTGACCAAGAGAGCAGCAATGCAGACTGGGTCACGAAGAAGCTCAGTGAAATCAATTATGAAGACCACAAGTTGAAGTCAGGGACTAACACCAAGAGGGCCCTCCAGGCAGTGTACAGCATGATGAGTTGGCCAGAGGACATCCCTCCTGAAGGCTGGAACCGCACCCGCCATGTCATCATCCTCATGACCGATGGATTGCACAACATGGGCGGGGACCCAATTACTGTCATTGATGAGATCCGGGACTTGTTATACATCGGCAAGGATCGTAAAAACCCGAGGGAGGATTATCTGGATGTCTATGTGTTTGGGGTTGGACCTTTGGTGGACCAAGTGAACATCAATGCTTTGGCTTCCAAGAAAGACAATGAGCAACATGTGTTCAAAGTCAAGGATATGGAAAACCTGGAAGACGTTTTCTTCCAAATGATTGATGAAAGCCAGTCTCTGAGTCTCTGTGGCATGGTTTGGGAACACACGACGGGTACCGATTACCACAAGCAACCATGGCAGGCCAAGATCTCAGTCACTCGCCCTTCGAAGGGACATGAGAGCTGTATGGGGGCTGTGGTGTCTGAGTACTTTGTGCTGACAGCAGCACATTGTTTTACTGTGGACGACAAGGAACACTCGATCAAGGTCAGCGTGGGGAAGAAGCGGGACCTGGAGATAGAAAAAGTCCTATTTCACCCCGACTACAACATTAGCGGGAAAAAAGAAGCAGGAATTCCTGAATTTTATGACTATGACGTTGCCCTGATCAAGCTCAAGAATAAGTTGAATTATGACCCGACTATCAGGCCCATTTGTCTCCCCTGCACCGAGGGAACAACTCGAGCTTTGAGGCTTCCTCCAACTACCACTTGCCAGCAACAGAAGGAAGAGCTGCTCCCTGCACAGGATATCAAAGCTCTGTTTGTGTCTGAGGAGGAGAAGAAGCTGACTCGGAAGGAGGTCTACATCAAGAATGGGGATAAGAAAGGCAGCTGTGAGAGAGATGCTCAATATGCCCCAGGCTATGACAAAGTCAAGGACATCTCCGAGGTGGTCACCCCTCGGTTCCTTTGTACTGGAGGAGTGAGTCCCTATGCTGACCCCAATACTTGCAGAGGTGATTCTGGCGGCCCCTTGATAGTTCACAAGAGAAGTCGTTTTATTCAAGTTGGTGTCATCAGCTGGGGAGTAGTGGATGTCTGCAAAAACCAGAAGCGGCAAAAGCAGGTACCTGCTCACGCCCGAGACTTTCACGTCAACCTCTTCCAAGTGCTGCCCTGGCTGAAGGAGAAACTCCAAGATGAGGATTTGGGTTTTCTC >gi|544428919|ref|XM_005553440.1|PREDICTED:Macaca fascicularis complement factor B (CFB),transcript variant X1, mRNA SEQ ID NO: 7ATTTCTGGTCCCTAAGTGGGTGGTCTGGGCTTGTTGGGGAGGAGCTGAGGCCAGAAGGAGGTACTGAAGGGGAGAGTCCTGGACCTTGGGCAGCAAAGGGTGGGACTTCTGCAGTTTCTGCTTCCTTGACTGGCAGCTCAGCGGGGCCCTCCCGCTTGGATGTTCCGGGAAAGTGATGAGGGTAGGACAGGCGGGGCAAGCTGCAGGTGCCAGAACACAGATTGCATAAAAGGCCGGGAGCTGGTGGGGGGCAGGGGAAGGGAATGTGACCAGGTCTAGGTCTGGAGTTTCAGCTTGGACACTGAGCTAAGTAGACAAGCAAAACAAGCCAGGACACGCCATCCTGCCCCAGGCCCAGCTTCTCTCCTGCCTTCTAACGCCATGGGGAGCAGTCTCAGCCCCCAGCTCTACCTGATGCCCTTCATCTTGGGCCTCTTATCTGCAGGTGTGACCACCACTCCATTGTCTTCGGCCCAGCCTCAAGGATCCTGCTCTCTGGAGGGGGTAGAGATCAAAGGTGGCTCCTTCCGACTTCTCCAAGAGGGCCAGGCACTGGAATACGTGTGTCCTTCTGGCTTCTACCCGTACCCTGTGCAGACACGTACCTGCAGATCCACGGGGTCCTGGAGCACCCTGCAGACTCAAGATCGAAAAACTGTCAAGAAGGCAGAGTGCAGAGCAATCCGCTGTCCACGACCACAGGACTTCGAGAACGGGGAATACCGGCCCCGGTCTCCCTACTACAATGTGAGTGATGAGATCTCTTTCCACTGCTATGACGGTTACACTCTCCGGGGCTCTGCCAATCGCACCTGCCAAGTGAATGGCCGGTGGAGTGGGCAGACAGCGATCTGTGACAACGGAGCGGGGTACTGCTCCAACCCAGGCATCCCCATTGGCACAAGGAAGGTGGGCAGCCGGTACCGCCTTGAAGACAGCGTCACCTACCACTGCAGCCGGGGGCTTACCCTGCGTGGCTCCCAGCGGCGAACGTGTCAGGAAGGTGGCTCTTGGAGCGGGACGGAGCCTTCCTGCCAAGACTCCTTCATGTACGACACCCCTCAAGAGGTGGCCGAAGCTTTCCTGTCTTCCCTGACGGAGACCATAGAAGGAGTCGATGCCGAGGATGGGCACAGCCCAGGGGAACAACAGAAGCGGAGGATCATCCTAGACCCTTCAGGCTCCATGAACATCTACCTGGTGCTAGATGGATCAGACAGCATTGGGGCCGGCAACTTCACAGGAGCCAAAAAGTGTCTAGTCAACTTAATTGAGAAGGTGGCAAGTTATGGTGTGAAGCCAAGATATGCTCTAGTGACATATGCCACATACCCCAGAATTTGGGTCAAAGTGTCTGACCAAGAGAGCAGCAATGCAGACTGGGTCACGAAGAAGCTCAGTGAAATCAATTATGAAGACCACAAGTTGAAGTCAGGGACTAACACCAAGAGGGCCCTCCAGGCAGTGTACAGCATGATGAGTTGGCCAGAGGACATCCCTCCTGAAGGCTGGAACCGCACCCGCCATGTCATCATCCTCATGACCGATGGATTGCACAACATGGGCGGGGACCCAATTACTGTCATTGATGAGATCCGGGACTTGTTATACATCGGCAAGGATCGCAAAAACCCGAGGGAGGATTATCTGGATGTCTATGTGTTTGGGGTTGGACCTTTGGTGGACCAAGTGAACATCAATGCTTTGGCTTCCAAGAAAGACAATGAGCAACATGTGTTCAAAGTCAAGGATATGGAAAACCTGGAAGACGTTTTCTTCCAAATGATTGATGAAAGCCAGTCTCTGAGTCTCTGTGGCATGGTTTGGGAACACACGACGGGTACCGATTACCACAAGCAACCATGGCAGGCCAAGATCTCAGTCACTCGCCCTTCGAAGGGACATGAGAGCTGTATGGGGGCTGTGGTGTCTGAGTACTTTGTGCTGACAGCAGCACATTGTTTTACTGTGGACGACAAGGAACACTCGATCAAGGTCAGCGTGGGGAAGAAGCGGGACCTGGAGATAGAAAAAGTCCTATTICACCCCGACTACAACATTAGCGGGAAAAAAGAAGCAGGAATTCCTGAATTTTATGACTATGACGTTGCCCTGATCAAGCTCAAGAAAAAGTTGAATTATGACCCGACTATCAGGCCCATTTGTCTCCCCTGTACCGAGGGAACAACTCGAGCTTTGAGGCTTCCTCCAACTACCACTTGCCAGCAACAGAAGGAAGAGCTGCTCCCTGCACAGGATATCAAAGCTCTGTTTGTGTCTGAGGAGGAGAAGAAGCTGACTCGGAAGGAGGTCTACATCAAGAATGGGGATAAGAAAGGCAGCTGTGAGAGAGATGCTCAATATGCCCCAGGCTATGACAAAGTCAAGGACATCTCGGAGGTGGTCACCCCTCGGTTCCTTTGTACTGGAGGAGTGAGTCCCTATGCTGACCCCAATACTTGCAGAGGTGATTCTGGCGGCCCCTTGATAGTTCACAAGAGAAGTCGTTTCATTCAAGTTGGTGTCATCAGCTGGGGAGTAGTGGATGTCTGCAAAAACCAGAAGCGGCAAAAGCAGGTACCTGCTCACGCCCGAGACTTTCACGTCAACCTCTTCCAAGTGCTGCCCTGGCTGAAGGAGAAACTCCAAGATGAGGATTTGGGTTTTCTCTAAGGGGTTTCCTGCTGGACAGGGGCGCGGGATTGAATTAAAACAGCTGCGACAACAReverse Complement of SEQ ID NO: 1 SEQ ID NO: 8TGTTGTCGCAGCTGTTTTAATTCAATCCCACGCCCCTGTCCAGCAGGAAACCCCTTATAGAAAACCCAAATCCTCATCTTGGAGTTTCTCCTTCAGCCAGGGCAGCACTTGAAAGAGGTTGATGTGAAAGTCTCGGGCGTGAGCAGGTACCTGCTTTTGCCGCTTCTGGTTTTTGCAGACATCCACTACTCCCCAGCTGATTACACCAACTTGAATGAAACGACTTCTCTTGTGAACTATCAAGGGGCCGCCAGAATCACCTCTGCAAGTATTGGGGTCAGCATAGGGACTCACTCCTCCAGTACAAAGGAACCGAGGGGTGACCACCTCTGAGATGTCCTTGACTTTGTCATAGCCTGGGGCATATTGAGCATCTCTCTCACAGCTGCCTTTCTTATCCCCATTCTTGATGTAGACCTCCTTCCGAGTCAGCTTTTTCTCCTCCTCAGACACAAACAGAGCTTTGATATCCTGTGCAGGGAGCAGCTCTTCCTTTTGTTGCTGGCAAGTGGTAGTTGGAGGAAGCCTCAAAGCTCGAGTTGTTCCCTCGGTGCAGGGGAGACAAATGGGCCTGATAGTCTGGCCATATTTCAGCTTATTCTTGAGCTTGATCAGGGCAACGTCATAGTCATAAAATTCAGGAATTCCTGCTTCTTTTTTCCCATTAATGTTGTAGTTGGGGTGAAATAGGACTACTTCTATCTCCAGGTCCCGCTTCTCCCCTCCTACGCTGACCTTGATTGAGTGTTCCTTGTCATCCACAGTGAAACAATGTGCTGCTGTCAGCACAAAGTACTCAGACACCACAGCCCCCATACAGCTCTCGTGTCCCTTTGAAGGGCGAATGACTGAGATCTTGGCCTGCCATGGTTGCTTGTGGTAATCGGTACCCTTCCTGTGTTCCCAAACCATGCCACAGAGACTCAGAGACTGGCTTTCATCGATCATTTGGTAGAAAACATCTTCCAGGTTTTCCATATCCTTGACTTTGAACACATGTTGCTCATTGTCTTTCTTGGAAGCCAAAGCATTGATGTTCACTTGGTTCACCAAAGGCCCGACCCCAAACACATAGACATCCAGATAATCCTCCCTTGGGTTTTTGCGATCCTTGCCAATGTATAGCAAGTCCCGGATCTCATCAATGACAGTAATTGGGTCCCCGCCCATGTTGTGCAATCCATCAGTCATGAGGATGATGACATGGCGGGTGCGGTTCCAGCCTTCAGGAGGGACGTCATCTGGCCAGCTCATCATGCTGTACACTGCCTGGAGGGCCTTCTTGGTGTTAGTCCCTGACTTCAACTTGTGGTCTTCATAATTGATTTCATTGAGCTGCTTCGTGACCCAGTCTGCATTACTGCTGTCTGCTTCAGACACTTTGACCCAAATTTTGGGGTATGTGGCATATGTCACTAGACCATATCTTGGCTTCACACCATAACTTGCCACCTTCTCAATTAAGTTGACTAGACACTTTTTGGCTCCTGTGAAGTTGCTGGCCCCAATGCTGTCTGATCCATCTAGCACCAGGTAGATGTTCATGGAGCCTGAAGGGTCCAGGACGATCTTCCGCTTCTGTTGTTCCCCTGGGCCGTGCCCATCCTCAGCATCGACTCCTTCTATGGTCTCTGTCAGGGAAGACAGGAAAGCTTCGGCCACCTCTTGAGGGGTGTCGTACATGAAGGAGTCTTGGCAGGAAGGCTCCGTCCCGCTCCAAGAGCCACCTTCCTGACACGTTCGCCGCTGGGAGCCACGCAGGGTAAGCCCCCGGCTGCAGTGGTAGGTGACGCTGTCTTCAAGGCGGTACTGGCTGCCCACCTTCCTTGTGCCAATGGGGATGCCCGGGTTGGAGCAGTACCCCGCTCCGTTGTCACAGATCGCTGTCTGCCCACTCCATCGGCCATTCACTTGGCAGGTGCGATTGGCAGAGCCCCGGAGAGTGTAACCGTCATAGCAGTGGAAAGAGATCTCATCACTCACATTGTAGTAGGGAGACCGGGGCCAGTATTCCCCGTTCTCGAAGTCGTGTGGTCTTGGACAGTGGATTGCTCTGCACTCTGCCTTCCTGACAGTCTTTTGGTCTTGAGTCTTCAGGGTGCTCCAGGACCCCGTAGATCTGCAGGTACGTGTCTGCACAGGGTACGGGTAGAAGCCAGAAGGACACACGTACTCCAGTGCCTGGCCCTCTTGGAGAAGTCGGAAGGAGCCGCCTTTGATCTCTACCCCCTCCAGAGAGCAGGATCCCTGGGGCCGGGCCAAAGACCATGGAGTGGTGGTCACACCTCCAGACAAGAGGCCCAAGATAAAGGGCATCAGGCAGAGTTGGGGGCTGAGATTGCTCCCCATGGCGTTGGAAGGCAGGAGAGAAGCTGGGCCTGGGGCAGGATGGTGTGTCCTGGCTTGCTTTGCTTGTCTGCTTGGCTCAGTGTCCAAGCTGAAACTCCAGACCTAGACCTGGTCACATTCCCTTCCC Reverse Complement of SEQ ID NO: 2SEQ ID NO: 9TTTTTTTTTTTTTTTTTTTTTTGTTATTGTAACTGCTTTAATCTGTCCTCACACTCTCCCTGCAGGAAGCTCTTTATAGAAAACCCAAATCCTCATCTTTGAGCTTGTCCTTTAGCCAGGGCAGCACCTGGAAGAGGTTGATGTGGAAGTCCCGGGCATAAGAGGGTACCAGCTGTTGCCGCCTCTGGTCTCTGCAGACATCTACTACTCCCCAGCTAATCACACCAACTTGAATGAAGCGGCTTCTCTTGTGAACAATGAGAGGGCCCCCGGAATCTCCTTTGCATGTGTTGGGGTCAGCATAGGGATCCACCCCTCCTGTGCAGAGGAACCGTGGAGTGACCACCTCAGAGGCATCTTTGACCTTCTCATAGCCTTGGGCCTTTGTAGCATCTCTCTCACAACTGGCTTTCTTGTCCCCATTCTTGATGTACACCTCCTTCCGAGTCAGGCTCTTCCCTTGCTCAGATACAAACAGAGCTTTGACATCCTTCACAGGGAGCAACTGTTCCTTGTGCTGCTTGCAGGTGGCTGTCTGAGGAAGCCTCAAGGCTCGTGTGGTTCCCTCCGTGCAGGGGAGACAGATGGGCCTGAGAGTCTGGCCATACTTGAGCTTGTTCTTGAGCTTGACTAGGGCCACATCATAATCATAGAACTCAGGGATCCCTTCTGCCTTTTTCCCATTAATATTGTATTTGGGGTGGAACAGGACCTCTTCAATCTCCAGGTCCCGCCTCTGACCCCCCACGCTGACCTTGATGGAATGTTTCTGATCATCCACCATGAAGCAGTGCGCTGCTGTCAGCACGAAGTACTCAGACACCACGGCCCCCATACAGGTCTCATGTCCTTTCAGAGGGCGAGTGACTGAGATCTTGGCTTGCCATGGTTGCTTATGATAATCGTTGCCTTTTTTATGCTCCCACACCATGCCACAGAGACTCAGAGATTTGGTTTCATCAATCATTTGGTAGAAAACATTCTCCAGGTCTTCCATATCCTTGACTTTAAACACATGATGCTCATTGTCCTTTTTGGAAGCTAAGGCATTGATGTTCACGGAGTCCACCAGAGGCCCGACCCCAAACACATACACATCCAGGTAATCCTCCCTGGGATTTTTGGGATCCCTGCCGATGTCCAGCAAGGCTCGGATGTCCTGAATGACAGTGACAGGGTTTCCACCCATGTTGTGCAAGCCATCAGTCATAATGATGATGACATGGCGGGTTCTATTCCAGCCTTCAGGCGGGGCATCCCCTGCCCAGCTCATCATGCTATACACAGCCTGGAGAGCCCTCTTGGTGTTGGTCCCTGACTTCAGCTTGTGGTCTTCATAACTGATTTGGTTGAGCTTCTCTGTGACCCAGTCGGCATCGCTACTCCTCTCATCAGACACTCTGACCAACACTTTGGGGACTGTAGCATATGTCAGGAGACCATATCGTGGCCTCACCCCGTAACTCGCCACCTTCTCAATCAAGTTGGTGAGGCACCGCTTAGCCCCTGTGAAGTTGCTGCTTCCGATGCTGTCTGATCCATCTAGCACCAGGTAGATATTCATGGAGCCCGAGGGGTCTAGGACAATCTTCCTCTTCTGCTGTTCTCCTGGGCTGTGCCCATCCTCAGCATCGGCTCCTTCGATGGTCTCTGTCAGGGAGGATAGGAATGCTTCGGCCACTTCTTGAGGGCTGTCATACATGAAGGAATCTTGGCAGGAAGGCTCTGTCCCACTCCATGAGCCACCTTCTTGACACTTTCGCTTCTGGGAGCCACGCAGGACAAGTCCCCGGCTGCAGTGGTAAGTAACAATGTCTTCAAGGCGGTATTGGCTACCCACCTTCCTTGTCCCAATAGGAATACCGGGATTGGGACAGTATCCAGCTCCATCATCACAAATTGCTGTTTGCCCATCCCACCGGCCATTCTCTTGGCAGGTGCGATTAGCAGAGCCCCGGAGAACGTAACCATCATAGCATTGAAAAGAAATCTGGTCACTCAGGTTGTAGAAGGGGGACCGGGGCCAGAATTCCCCATTTTCAAAGTCCTGCGGTCGTGGGCAGCGTATTGCTCTGCATTCCGCCTTCTGGACAATCTTTTGGTCTCGGGTCTGCAGGTCGCTCCAGGAGCCTGTGGATCTGCAGGTTCGAGTCTGCACGGGGTATGGGTAGAAGCCAGAGGGACATAGGTACTCCAGGGCCTGACCGCCTTGGAGAAGTTGAAAGGAGCCGCCTTTGATCTCTACTCCCTCCAGAGAGCAGGAGACTTGGGGCCGGGCCTCAAGCACTGGAGTTGCGCTCACACCTCCAGAGGAGAAGCCTAAGACCAAGAGGACGAGGCAGAGCTGGGGGCTCTCCATTGTCATGGAAAGCACCAGAGAGTCCAGTGGCCCAAAGCCCTCTCCAGGTCACTCTGGGTGCCTATTTGACCTCCTGTCCAAACTGAAACCCAAATGCTGGATCTGGCTACACCCCTTTCCCACTGTCCCCTAACCTTTTATGCAATCTGCTCTGGCATCTGCAGCCTGGCCAGCCTACCCTGCGCACGCAGCCACTGCTTCCCGGAACATTCATGAAGGAGGGCCCCAGCTGAACTGCCAACTGAGGAAACAGAAACTATGTGAGCCCCACCCTTGACTGACCAAGGGCCAGAACTTCCCTTTCAAAGCTCCCTCTTAGCCCCCAGACCCCTCCTAACAAGCACAGACCACCACCTCCACTACCCCCAATTTAAAAGATCAGTCTTTCCATGGACTGTGTGATGGAGC Reverse Complement of SEQ ID NO: 3SEQ ID NO: 10TTTTTTTTTTTTTTTTTTTTTTGTTATTGTAACTGCTTTAATCTGTCCTCACACTCTCCCTGCAGGAAGCTCTTTATAGAAAACCCAAATCCTCATCTTTGAGCTTGTCCTTTAGCCAGGGCAGCACCTGGAAGAGGTTGATGTGGAAGTCCCGGGCATAAGAGGGTACCAGCTGTTGCCGCCTCTGGTCTCTGCAGACATCTACTACTCCCCAGCTAATCACACCAACTTGAATGAAGCGGCTTCTCTTGTGAACAATGAGAGGGCCCCCGGAATCTCCTTTGCATGTGTTGGGGTCAGCATAGGGATCCACCCCTCCTGTGCAGAGGAACCGTGGAGTGACCACCTCAGAGGCATCTTTGACCTTCTCATAGCCTTGGGCCTTTGTAGCATCTCTCTCACAACTGGCTTGTCCCCATTCTTGATGTACACCTCCTTCCGAGTCAGGCTCTTCCCTTGCTCAGATACAAACAGAGCTTTGACATCCTTCACAGGGAGCAACTGTTCCTTGTGCTGCTTGCAGGTGGCTGTCTGAGGAAGCCTCAAGGCTCGTGTGGTTCCCTCCGTGCAGGGGAGACAGATGGGCCTGAGAGTCTGGCCATACTTGAGCTTGTTCTTGAGCTTGACTAGGGCCACATCATAATCATAGAACTCAGGGATCCCTTCTGCCTTTTTCCCATTAATATTGTATTTGGGGTGGAACAGGACCTCTTCAATCTCCAGGTCCCGCCTCTGACCCCCCACGCTGACCTTGATGGAATGTTTCTGATCATCCACCATGAAGCAGTGCGCTGCTGTCAGCACGAAGTACTCAGACACCACGGCCCCCATACAGGTCTCATGTCCTTTCAGAGGGCGAGTGACTGAGATCTTGGCTTGCCATGGTTGCTTATGATAATCGTTGCCTTTTTTATGCTCCCACACCATGCCACAGAGACTCAGAGATTIGGTTTCATCAATCATTTGGTAGAAAACATTCTCCAGGTCTTCCATATCCTTGACTTTAAACACATGATGCTCATTGTCCTTTTTGGAAGCTAAGGCATTGATGTTCACGGAGTCCACCAGAGGCCCGACCCCAAACACATACACATCCAGGTAATCCTCCCTGGGATTTTTGGGATCCCTGCCGATGTCCAGCAAGGCTCGGATGTCCTGAATGACAGTGACAGGGTTTCCACCCATGTTGTGCAAGCCATCAGTCATAATGATGATGACATGGCGGGTTCTATTCCAGCCTTCAGGCGGGGCATCCCCTGCCCAGCTCATCATGCTATACACAGCCTGGAGAGCCCTCTTGGTGTTGGTCCCTGACTTCAGCTTGTGGTCTTCATAACTGATTTGGTTGAGCTTCTCTGTGACCCAGTCGGCATCGCTACTCCTCTCATCAGACACTCTGACCAACACTTTGGGGACTGTAGCATATGTCAGGAGACCATATCGTGGCCTCACCCCGTAACTCGCCACCTTCTCAATCAAGTTGGTGAGGCACCGCTTAGCCCCTGTGAAGTTGCTGCTTCCGATGCTGTCTGATCCATCTAGCACCAGGTAGATATTCATGGAGCCCGAGGGGTCTAGGACAATCTTCCTCTTCTGCTGTTCTCCTGGGCTGTGCCCATCCTCAGCATCGGCTCCTTCGATGGTCTCTGTCAGGGAGGATAGGAATGCTTCGGCCACTTCTTGAGGGCTGTCATACATGAAGGAATCTTGGCAGGAAGGCTCTGTCCCACTCCATGAGCCACCTTCTTGACACTTTCGCTTCTGGGAGCCACGCAGGACAAGTCCCCGGCTGCAGTGGTAAGTAACAATGTCTTCAAGGCGGTATTGGCTACCCACCTTCCTTGTCCCAATAGGAATACCGGGATTGGGACAGTATCCAGCTCCATCATCACAAATTGCTGTTTGCCCATCCCACCGGCCATTCTCTTGGCAGGTGCGATTAGCAGAGCCCCGGAGAACGTAACCATCATAGCATTGAAAAGAAATCTGGTCACTCAGGTTGTAGAAGGGGGACCGGGGCCAGAATTCCCCATTTTCAAAGTCCTGCGGTCGTGGGCAGCGTATTGCTCTGCATTCCGCCTTCTGGACAATCTTTTGGTCTCGGGTCTGCAGGTCGCTCCAGGAGCCTGTGGATCTGCAGGTTCGAGTCTGCACGGGGTATGGGTAGAAGCCAGAGGGACATAGGTACTCCAGGGCCTGACCGCCTTGGAGAAGTTGAAAGGAGCCGCCTTTGATCTCTACTCCCTCCAGAGAGCAGGAGACTTGGGGCCGGGCCTCAAGCACTGGAGTTGCGCTCACACCTCCAGAGGAGAAGCCTAAGACCAAGAGGACGAGGCAGAGCTGGGGGCTCTCCATTGTCATGGAAAGCACCAGAGAGTCCAGTGGCCCAAAGCCCTCTCCAGGTCACTCTGGGTGCCTATTTGACCTCCTGTCCAAACTGAAACCCAAATGCTGGATCTGGCTACACCCCTTTCCCACTGTCCCCTAACCTTTTATGCAATCTGCTCTGGCATCTGCAGCCTGGCCAGCCTACCCTGCGCACGCAGCCACTGCTTCCCGGAACATTCATGAAGGAGGGCCCCAGCTGAACTGCCAACTGAGGAAACAGAAACTATGTGAGCCCCACCCTTGACTGACCAAGGGCCAGAACTTCCCTTTCAAAGCTCCCTCTTAGCCCCCAGACCCCTCCTAACAAGCACAGACCACCACCTCCACTACCCCCAATTTAAAAGATCAGTCTTTCCATGGACTGTGTGATGGAGC Reverse Complement of SEQ ID NO: 4SEQ ID NO: 11TTTTTTTTTTTTTTTTGTATTTGTATTGTAGCTGCTTTAATCTGCCCTCACCCTCCCAGCAGGAAGCTCCTTATAAGAAACCCAAGTCCTCGTCTTTGAGCTTCTCCTTTAGCCAGGGCAGCACCTGGAAGAGATTGATGTGGAAGTCCCGGGCATAGGAGGGCACCAACTGTTGCCGCCTCGGGTCTTTGCAGACATCCACTACTCCCCAGCTGATCACACCAACTTGAATGAAGCGGCTTCTCTTGTGAACAATGAGAGGGCCCCCGGAGTCTCCTTTGCATGTGTTGGGGTCAGCATAGGGATCTACCCCTCCGGTGCACAGGAACCTGGGGGTGACCACCTCAGAGGCAACTTTGACCTTCTCATAGCCTTGGGCCTTTGTAGCATCTCTCTCACAACTGGCTTTCTTTTCCCCATTCTTGATGTACACCTCCTTCCGGGTCAGCTTCTTCCCTTCCTCGGATACAAACAGAGCTTTGACGTCCTTCATAGGGAGCAACTCTTCCTTGTGCTGTTTGCAGGTGGCTGTCTGAGGAAGCCGCAAGGCTCGGGTGGTTCCCTCTGTGCAGGGGAGACAGATGGGCCTGAGAGTCTGGCTGTACTTCAGCTTGGTCTTGAGCTTGATGAGGGCAACATCATAGTCATAGAACTCAGAGATTCCTTCTGCCTTTTTCCCATTGATGTCGTAATTAGGGTGGAACAGGACCTCTTCAATCTCCAGGTCCCGCCTTTTCCCCTCCACGTTGACCTTGATGGAGTGTTTCTGATCTTCCACTGTGAAGCAATGCGCTGCTGTCAGCACGAAGTACTCGGACACCACGGCCCCCATACAGTTCTCATGTCCTTTCAGAGGACGAGTGACTGAGATCTTGGCTTGCCATGGTTGCTTGTAATAATCACCGCCTTTCTGATGCTCCCACACCATGCCACAGAGACCCAGAGATTTGGTTTCATCGATCATTTTGTAGAAGACGTTCTCCAGATCCTCCATGTCCTTGACCTTGAACACATGCTGCTCATTGTTCTTTTTGGAAGCCAAGGCATTGATGTTCACAGGGTCCACCAGAGGCCCGACCCCAAACACATACACATCCAAATAATCCTCCCGGGGATTTTTGCGATCCCTGCCAATATCCAGCAAGTCTCGGATGTCCTCAATGACAGTGACAGGGTCTCCACCCATGTTGTGCAAGCCATCAGTCATGATGATGATGACGTGGCGGGTTCGATTCCAGCCTTCAGGCGGAGCATCCCCTGGCCAGCTCATCATGCTGTATACAGCCTGGAGAGCCTTCTTGGTGTTGGTCCCTGACTTCAGCTTGTGGTCTTCATAACTGATTTGGTTGAGCTTCTCTGTGACCCAGTCGGCATCACTACTCCTCTCCTCAGACACTCTGACCAAGACTTTGGGGACTGTGGCATATGTCACTAGACCGTATCTTGGCTTCACCCCATAACTCGCCACCTTCTCAATCAAGTTGGCGAGACACCGCTTGGCCCCTGTGAAGTTGCTGGCCCCGATGCTGTCGGATCCATCCAGCACCATGTAGATATTCATGGAGCCCGAGGGGTCCAGGATAATCTTCCTCTTCTGCTGTTCCCCTGGGCTGTGCCCATCCTCCGCATCTGCTCCTTCGATGGTCTCTGTCAGGGAGGATAGAAATGCTTCGGCCACCTCTTGAGGGCTGTCGTACATGAAGGAATCTTGGCAGGAAGGCTCTGTCCCACTCCACGAGCCACCTTCCTGGCACCTTCGCTGCTGGGAGCCACGTAGGACAAGTCCCCGACTACAGTGGTAAGTGACAGTGTCTTCAAGACGGTACTGGCTTCCCACCTTCCTTGTCCCAATAGGAATACCCGGGTTGGGACAGTATCCCGCTCCATCATCACAGATTGCTGTTTGCCCATCCCACCGGCCATTCTCTTGGCAGGTGCGATTAGCAGAGCCCCGGAGAGTGTAGCCATCATAGCATTGAAAAGAAATCTGATCACTCAGGTTGTAGTAGGGGGACCGGGGCCAGAACTCCCCATTTTCAAAGTCCTGTGGTCGTGGGCAGCGTATTGCTCTGCATTCTGCCTTCTTGACAATCTTTTGGTCCCGGGTCTGGAGGACACTCCAGGAGCCTGTGGATTTGCAGGTTCGAGTCTGCACAGGGTATGGGTAGAAGCCAGAGGGACACAGGTACTCCAGGGCCTGACCGTCTTGGAGAAGTTGGAAGGAGCCGCCTTTGATCTCTACTCCCTCCAGAGAGCAAGAGACCTGGGGCCGGGCCTCAAGCACTGGAGTTGCGCTCACACCTCCGGAGGAGAGGCCTAAGACCAAGAGGACTAAGCAGAGCTGGGGACCCTCCATTGTCATGGAAAGCCCGTGGCCAAAAGCCCTTTCCAGTTCACTCTGGGTGGCTACTTGACTTGCTGTCCAGACTGAAACTCCAGTGCTGGATCTGGCTACACCCCTTTCCCACCGGCCCCTAACCTTTTATGCAATCTGCTCTGGCACCTGCAGCCTGGCCAACCTACCCTGCACAGACGCTGCTTCCCGGAACATTCATGAAGGAGGGCCCCTGCTGReverse Complement of SEQ ID NO: 5 SEQ ID NO: 12ACCCCTTATAGAAAACCCAAATCCTCATCTTGGAGTTTCTCCTTCAGCCAGGGCAGCACTTGAAAGAGGTTGATGTGAAAGTCTCGGGCGTGAGCAGGTACCTGCTTTTGCCGCTTCTGGTTTTTGCAGACATCCACTACTCCCCAGCTGATTACACCAACTTGAATGAAACGACTTCTTTTGTGAACTATCAAGGGGCCGCCAGAATCACCTCTGCAAGTATTGGGGTCAGCATAGGGACTCACTCCTCCAGTACAAAGGAACCGAGGGGTGACCACCTCTGAGATGTCCTTGACTTTGTCATAGCCTGGGGCATATTGAGCATCTCTCTCACAGCTGCCTTTCTTATCCCCATTCTTGATGTAGACCTCCTTCCGAGTCAGCTTTTTCTCCTCCTCAGACACAAACAGAGCTTTGATATCCTGTGCAGGGAGCAGCTCTTCCTTTTGTTGCTGGCAAGTGGTAGTTGGAGGAAGCCTCAAAGCTCGAGTTGTTCCCTCGGTGCAGGGGAGACAAATGGGCCTGATAGTCTGGCCATATTTCAGCTTATTCTTGAGCTTGATCAGGGCAACGTCATAGTCATAAAATTCAGGAATTCCTGCTGCTTTTTTCCCATTAATGTTGTAGTTGGGGTGAAATAGGACTACTTCTATCTCCAGGTCCCGCTTCTCCCCTCCTACGCTGACCTTGATTGAGTGTTCCTTGTCATCCACAGTGAAACAGTGTGCTGCTGTCAGCACAAAGTACTCAGACACCACAGCCCCCATACAGCTCTCGTGTCCCTTTGAAGGGCGAATGACTGAGATCTTGGCTTGCCATGGTTGCTTGTGGTAATCGGTACCCTTCCTGTGTTCCCAAACCATGCCACAGAGACTCAGAGACTGGCTTTCATCAATCATTTGGTAGAAAACATCTTCCAGGTTTTCCATATCCTTGACTTTGAACACATGTTGCTCATTGTCTTTCTTGGAAGCCAAAGCATTGATGTTCACTTGGTTCACCAAAGGCCCGACCCCAAACACATAGACATCCAGATAATCCTCCCTTGGGTTTTTGCGATCCTTGCCAATGTATAGCAAGTCCCGGATCTCATCAATGACAGTAATTGGGTCCCCGCCCATGTTGTGCAATCCATCAGTCATGAGGATGATGACATGGCGGGTGCGGTTCCAGCCTTCAGGAGGGATGTCATCTGGCCAGCTCATCATGCTGTACACTGCCTGGAGGGCCTTCTTGGTGTTAGTCCCTGACTTCAACTTGTGGTCTTCATAATTGATTTCATTGAGCTGCTTCGTGACCCAGTCTGCATTACTGCTGTCTGGATCAGACACTTTGACCCAAATTTTGGGGTGTGTGGCATATGTCACTAGACCATATCTTGGCTTCACACCATAACTTGCCACCTTCTCAATTAAGTTGACTAGACACTTTTTGGCTCCTGTGAAGTTGCTGGCCCCAATGCTGTCTGATCCATCTAGCACCAGGTAGATGTTCATGGAGCCTGAAGGGTCCAGGACGATCTTCCGCTTCTGTTGTTCCCCTGGGCCGTGCCCATCCTCAGCATCGACTCCTTCTATGGTCTCTGTCAGGGAAGACAGGAAAGCTTCGGCCACCTCTTGAGGGGTGTCGTACATGAAGGAGTCTTGGCAAGAAGGCTCCGTCCCGCTCCAAGAGCCACCTTCCTGACACGTTCGCCGCTGGGAGCCACGCAGGGTAAGCCCCCGGCTGCAGTGGTAGGTGACGCTGTCTTCAAGGCGGTACTGGCTGCCCACCTTCCTTGTGCCAATGGGGATGCCCGGGTTGGAGCAGTACCCCGCTCCGTTGTCACAGATCGCTGTCTGCCCACTCCACCGGCCATTCACTTGGCAGGTGCGATTGGCAGAGCCCCGGAGAGTGTAACCGTCATAGCAGTGGAAAGAGATCTCATCACTCACATTGTAGTAGGGAGACCGGGGCCAGTATTCCCCGTTCTCGAAGTCGTGTGGTCTTGGACAGTGGATTGCTCTGCACTCTGCCTTCCTGACAGTCTTTTGGACTTGAGTCTTCAGGGTGCTCCAGGACCCCGTAGATCTGCAGGTACGTGTCTGCACAGGGTACGGGTAGAAGCCAGAAGGACACACGTACTCCAGTGCCTGGCCCTCTTGGAGAAGTCGGAAGGAGCCGCCTTTGATCTCTACCCCCTCCAGAGAGCAGGATTCCTGGGGCTGGGCCAAAGGCCATGGAGTGGTGGTCACACCTCCAGACAAGAGGCCCAAGATGAAGGGCATCAGGCAGAGTTGGGGGCTGAGATTGCTCCCCATGGCGTTGGAAGGCAGGAGAGAAGCTGGGCCTGGG Reverse Complement of SEQ ID NO: 6SEQ ID NO: 13GAGAAAACCCAAATCCTCATCTTGGAGTTTCTCCTTCAGCCAGGGCAGCACTTGGAAGAGGTTGACGTGAAAGTCTCGGGCGTGAGCAGGTACCTGCTTTTGCCGCTTCTGGTTTTTGCAGACATCCACTACTCCCCAGCTGATGACACCAACTTGAATAAAACGACTTCTCTTGTGAACTATCAAGGGGCCGCCAGAATCACCTCTGCAAGTATTGGGGTCAGCATAGGGACTCACTCCTCCAGTACAAAGGAACCGAGGGGTGACCACCTCGGAGATGTCCTTGACTTTGTCATAGCCTGGGGCATATTGAGCATCTCTCTCACAGCTGCCTTTCTTATCCCCATTCTTGATGTAGACCTCCTTCCGAGTCAGCTTCTTCTCCTCCTCAGACACAAACAGAGCTTTGATATCCTGTGCAGGGAGCAGCTCTTCCTTCTGTTGCTGGCAAGTGGTAGTTGGAGGAAGCCTCAAAGCTCGAGTTGTTCCCTCGGTGCAGGGGAGACAAATGGGCCTGATAGTCGGGTCATAATTCAACTTATTCTTGAGCTTGATCAGGGCAACGTCATAGTCATAAAATTCAGGAATTCCTGCTTCTTTTTTCCCGCTAATGTTGTAGTCGGGGTGAAATAGGACTTTTTCTATCTCCAGGTCCCGCTTCTTCCCCACGCTGACCTTGATCGAGTGTTCCTTGTCGTCCACAGTAAAACAATGTGCTGCTGTCAGCACAAAGTACTCAGACACCACAGCCCCCATACAGCTCTCATGTCCCTTCGAAGGGCGAGTGACTGAGATCTTGGCCTGCCATGGTTGCTTGTGGTAATCGGTACCCGTCGTGTGTTCCCAAACCATGCCACAGAGACTCAGAGACTGGCTTTCATCAATCATTTGGAAGAAAACGTCTTCCAGGTTTTCCATATCCTTGACTTTGAACACATGTTGCTCATTGTCTTTCTTGGAAGCCAAAGCATTGATGTTCACTTGGTCCACCAAAGGTCCAACCCCAAACACATAGACATCCAGATAATCCTCCCTCGGGTTTTTACGATCCTTGCCGATGTATAACAAGTCCCGGATCTCATCAATGACAGTAATTGGGTCCCCGCCCATGTTGTGCAATCCATCGGTCATGAGGATGATGACATGGCGGGTGCGGTTCCAGCCTTCAGGAGGGATGTCCTCTGGCCAACTCATCATGCTGTACACTGCCTGGAGGGCCCTCTTGGTGTTAGTCCCTGACTTCAACTTGTGGTCTTCATAATTGATTTCACTGAGCTTCTTCGTGACCCAGTCTGCATTGCTGCTCTCTTGGTCAGACACTTTGACCCAAATTCTGGGGTATGTGGCATATGTCACTAGAGCATATCTTGGCTTCACACCATAACTTGCCACCTTCTCAATTAAGTTGACTAGACACTTTTTGGCTCCTGTGAAGTTGCCGGCCCCAATGCTGTCTGATCCATCTAGCACCAGGTAGATGTTCATGGAGCCTGAAGGGTCTAGGATGATCCTCCGCTTCTGTTGTTCCCCTGGGCTGTGCCCATCCTCGGCATCGACTCCTTCTATGGTCTCCGTCAGGGAAGACAGGAAAGCTTCGGCCACCTCTTGAGGGGTGTCGTACATGAAGGAGTCTTGGCAGGAAGGCTCCGTCCCGCTCCAAGAGCCACCTTCCTGACATGTTCGCCGCTGGGAGCCACGCAGGGTAAGCCCCCGGCTGCAGTGGTAGGTGACGCTGTCTTCAAGGCGGTACCGGCTGCCCACCTTCCTTGTGCCAATGGGGATGCCTGGGTTGGAGCAGTACCCCGCTCCGTTGTCACAGATCGCTGTCTGCCCACTCCACCGGCCATTCACTTGGCAGGTGCGATTGGCAGAGCCCCGGAGAGTGTAACCGTCATAGCAGTGGAAAGAGATCTCATCACTCACATTGTAGTAGGGAGACCGGGGCCGGTATTCCCCGTTCTCGAAGTCCTGTGGTCGTGGACAGCGGATTGCTCTGCACTCTGCCTTCTTGACAGTTTTTCGATCTTGAGTCTGCAGGGTGCTCCAGGACCCCGTGGATCTGCAGGTACGTGTCTGCACAGGGTACGGGTAGAAGCCAGAAGGACACACGTATTCCAGTGCCTGGCCCTCTTGGAGAAGTCGGAAGGAGCCACCTTTGATCTCTACCCCCTCCAGAGAGCAGGATCCTTGAGGCTGGGCCGAAGACAATGGAGTGGTGGTCACACCTGCAGATAAGAGGCCCAAGATGAAGGGCATCAGGTAGAGCTGGGGGCTGAGACTGCTCCCCATReverse Complement of SEQ ID NO: 7 SEQ ID NO: 14TGTTGTCGCAGCTGTTTTAATTCAATCCCGCGCCCCTGTCCAGCAGGAAACCCCTTAGAGAAAACCCAAATCCTCATCTTGGAGTTTCTCCTTCAGCCAGGGCAGCACTTGGAAGAGGTTGACGTGAAAGTCTCGGGCGTGAGCAGGTACCTGCTTTTGCCGCTTCTGGTTTTTGCAGACATCCACTACTCCCCAGCTGATGACACCAACTTGAATGAAACGACTTCTCTTGTGAACTATCAAGGGGCCGCCAGAATCACCTCTGCAAGTATTGGGGTCAGCATAGGGACTCACTCCTCCAGTACAAAGGAACCGAGGGGTGACCACCTCCGAGATGTCCTTGACTTTGTCATAGCCTGGGGCATATTGAGCATCTCTCTCACAGCTGCCTTTCTTATCCCCATTCTTGATGTAGACCTCCTTCCGAGTCAGCTTCTTCTCCTCCTCAGACACAAACAGAGCTTTGATATCCTGTGCAGGGAGCAGCTCTTCCTTCTGTTGCTGGCAAGTGGTAGTTGGAGGAAGCCTCAAAGCTCGAGTTGTTCCCTCGGTACAGGGGAGACAAATGGGCCTGATAGTCGGGTCATAATTCAACTTTTTCTTGAGCTTGATCAGGGCAACGTCATAGTCATAAAATTCAGGAATTCCTGCTTCTTTTTTCCCGCTAATGTTGTAGTCGGGGTGAAATAGGACTTTTTCTATCTCCAGGTCCCGCTTCTTCCCCACGCTGACCTTGATCGAGTGTTCCTTGTCGTCCACAGTAAAACAATGTGCTGCTGTCAGCACAAAGTACTCAGACACCACAGCCCCCATACAGCTCTCATGTCCCTTCGAAGGGCGAGTGACTGAGATCTTGGCCTGCCATGGTTGCTTGTGGTAATCGGTACCCGTCGTGTGTTCCCAAACCATGCCACAGAGACTCAGAGACTGGCTTTCATCAATCATTTGGAAGAAAACGTCTTCCAGGTTTTCCATATCCTTGACTTTGAACACATGTTGCTCATTGTCTTTCTTGGAAGCCAAAGCATTGATGTTCACTTGGTCCACCAAAGGTCCAACCCCAAACACATAGACATCCAGATAATCCTCCCTCGGGTTTTTGCGATCCTTGCCGATGTATAACAAGTCCCGGATCTCATCAATGACAGTAATTGGGTCCCCGCCCATGTTGTGCAATCCATCGGTCATGAGGATGATGACATGGCGGGTGCGGTTCCAGCCTTCAGGAGGGATGTCCTCTGGCCAACTCATCATGCTGTACACTGCCTGGAGGGCCCTCTTGGTGTTAGTCCCTGACTTCAACTTGTGGTCTTCATAATTGATTTCACTGAGCTTCTTCGTGACCCAGTCTGCATTGCTGCTCTCTTGGTCAGACACTTTGACCCAAATTCTGGGGTATGTGGCATATGTCACTAGAGCATATCTTGGCTTCACACCATAACTTGCCACCTTCTCAATTAAGTTGACTAGACACTTTTTGGCTCCTGTGAAGTTGCCGGCCCCAATGCTGTCTGATCCATCTAGCACCAGGTAGATGTTCATGGAGCCTGAAGGGTCTAGGATGATCCTCCGCTTCTGTTGTTCCCCTGGGCTGTGCCCATCCTCGGCATCGACTCCTTCTATGGTCTCCGTCAGGGAAGACAGGAAAGCTTCGGCCACCTCTTGAGGGGTGTCGTACATGAAGGAGTCTTGGCAGGAAGGCTCCGTCCCGCTCCAAGAGCCACCTTCCTGACACGTTCGCCGCTGGGAGCCACGCAGGGTAAGCCCCCGGCTGCAGTGGTAGGTGACGCTGTCTTCAAGGCGGTACCGGCTGCCCACCTTCCTTGTGCCAATGGGGATGCCTGGGTTGGAGCAGTACCCCGCTCCGTTGTCACAGATCGCTGTCTGCCCACTCCACCGGCCATTCACTTGGCAGGTGCGATTGGCAGAGCCCCGGAGAGTGTAACCGTCATAGCAGTGGAAAGAGATCTCATCACTCACATTGTAGTAGGGAGACCGGGGCCGGTATTCCCCGTTCTCGAAGTCCTGTGGTCGTGGACAGCGGATTGCTCTGCACTCTGCCTTCTTGACAGTTTTTCGATCTTGAGTCTGCAGGGTGCTCCAGGACCCCGTGGATCTGCAGGTACGTGTCTGCACAGGGTACGGGTAGAAGCCAGAAGGACACACGTATTCCAGTGCCTGGCCCTCTTGGAGAAGTCGGAAGGAGCCACCTTTGATCTCTACCCCCTCCAGAGAGCAGGATCCTTGAGGCTGGGCCGAAGACAATGGAGTGGTGGTCACACCTGCAGATAAGAGGCCCAAGATGAAGGGCATCAGGTAGAGCTGGGGGCTGAGACTGCTCCCCATGGCGTTAGAAGGCAGGAGAGAAGCTGGGCCTGGGGCAGGATGGCGTGTCCTGGCTTGTTTTGCTTGTCTACTTAGCTCAGTGTCCAAGCTGAAACTCCAGACCTAGACCTGGTCACATTCCCTTCCCCTGCCCCCCACCAGCTCCCGGCCTTTTATGCAATCTGTGTTCTGGCACCTGCAGCTTGCCCCGCCTGTCCTACCCTCATCACTTTCCCGGAACATCCAAGCGGGAGGGCCCCGCTGAGCTGCCAGTCAAGGAAGCAGAAACTGCAGAAGTCCCACCCTTTGCTGCCCAAGGTCCAGGACTCTCCCCTTCAGTACCTCCTTCTGGCCTCAGCTCCTCCCCAACAAGCCCAGACCACCCACTTAGGGACCAGAAAT

We claim:
 1. A double stranded ribonucleic acid (dsRNA) agent, or saltthereof, for inhibiting expression of complement factor B (CFB) in acell, wherein the dsRNA agent, or salt thereof, comprises a sense strandand an antisense strand forming a double stranded region, wherein thesense strand comprises a nucleotide sequence which differs by no morethan 4 nucleotides from the nucleotide sequence5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ ID NO:1271) and the antisensestrand comprises a nucleotide sequence which differs by no more than 4nucleotides from the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922).
 2. The dsRNAagent, or salt thereof, of claim 1, wherein the sense strand comprises anucleotide sequence which differs by no more than 3 nucleotides from thenucleotide sequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ ID NO:1271)and the antisense strand comprises a nucleotide sequence which differsby no more than 3 nucleotides from the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922).
 3. The dsRNAagent, or salt thereof, of claim 1, wherein the sense strand comprises anucleotide sequence which differs by no more than 2 nucleotides from thenucleotide sequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ ID NO:1271)and the antisense strand comprises a nucleotide sequence which differsby no more than 2 nucleotides from the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922).
 4. The dsRNAagent, or salt thereof, of claim 1, wherein the sense strand comprises anucleotide sequence which differs by no more than 1 nucleotide from thenucleotide sequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ ID NO:1271)and the antisense strand comprises a nucleotide sequence which differsby no more than 1 nucleotide from the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922).
 5. The dsRNAagent, or salt thereof, of claim 1, wherein the sense strand comprisesthe nucleotide sequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ IDNO:1271) and the antisense strand comprises the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922).
 6. The dsRNAagent, or salt thereof, of claim 1, wherein the sense strand consists ofthe nucleotide sequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ IDNO:1271) and the antisense strand consists of the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922).
 7. The dsRNAagent, or salt thereof, of claim 1, further comprising a ligand.
 8. ThedsRNA agent, or salt thereof, of claim 7, wherein the ligand isconjugated to the 3′ end of the sense strand of the dsRNA agent.
 9. ThedsRNA agent, or salt thereof, of claim 7, wherein the ligand is anN-acetylgalactosamine (GalNAc) derivative.
 10. The dsRNA agent, or saltthereof, of claim 9, wherein the ligand is conjugated to the dsRNA agentthrough a monovalent, bivalent, or trivalent linker.
 11. The dsRNAagent, or salt thereof, of claim 9, wherein the ligand is


12. The dsRNA agent, or salt thereof, of claim 11, wherein the dsRNAagent, or salt thereof, is conjugated to the ligand as shown in thefollowing schematic

wherein X is O or S.
 13. The dsRNA agent, or salt thereof, of claim 12,wherein the X is O.
 14. An isolated cell containing the dsRNA agent, orsalt thereof, of claim
 1. 15. A pharmaceutical composition comprisingthe dsRNA agent, or salt thereof, of claim
 1. 16. The pharmaceuticalcomposition of claim 15, wherein the dsRNA agent, or salt thereof, is inan unbuffered solution.
 17. The pharmaceutical composition of claim 16,wherein the unbuffered solution is saline or water.
 18. Thepharmaceutical composition of claim 15, wherein the dsRNA agent, or saltthereof, is in a buffer solution.
 19. The pharmaceutical composition ofclaim 18, wherein the buffer solution comprises acetate, citrate,prolamine, carbonate, or phosphate or any combination thereof.
 20. Thepharmaceutical composition of claim 19, wherein the buffer solution isphosphate buffered saline (PBS).
 21. A double stranded ribonucleic acid(dsRNA) agent, or salt thereof, for inhibiting expression of complementfactor B (CFB) in a cell, wherein the dsRNA agent, or salt thereof,comprises a sense strand and an antisense strand forming a doublestranded region, wherein the sense strand comprises the nucleotidesequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′ (SEQ ID NO:1271) and theantisense strand comprises the nucleotide sequence5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922), and wherein the3′-end of the sense strand is conjugated to a ligand as shown in thefollowing schematic

wherein X is O.
 22. An isolated cell containing the dsRNA agent, or saltthereof, of claim
 21. 23. A pharmaceutical composition comprising thedsRNA agent, or salt thereof, of claim
 21. 24. The pharmaceuticalcomposition of claim 23, wherein dsRNA agent, or salt thereof, is in anunbuffered solution.
 25. The pharmaceutical composition of claim 15,wherein the dsRNA agent, or salt thereof, is in a buffer solution.
 26. Adouble stranded ribonucleic acid (dsRNA) agent, or salt thereof, forinhibiting expression of complement factor B (CFB) in a cell, whereinthe dsRNA, or salt thereof, comprises a sense strand and an antisensestrand forming a double stranded region, wherein the sense strandconsists of the nucleotide sequence 5′-gsasauuccuGfAfAfuuuuaugacu-3′(SEQ ID NO:1271) and the antisense strand consists of the nucleotidesequence 5′-asdGsucdAudAaaaudTcAfggaauucscsu-3′ (SEQ ID NO:1922), andwherein the 3′-end of the sense strand is conjugated to a ligand asshown in the following schematic

wherein X is O.
 27. An isolated cell containing the dsRNA agent, or saltthereof, of claim
 26. 28. A pharmaceutical composition comprising thedsRNA agent, or salt thereof, of claim
 26. 29. The pharmaceuticalcomposition of claim 28, wherein dsRNA agent, or salt thereof, is in anunbuffered solution.
 30. The pharmaceutical composition of claim 28,wherein said dsRNA agent, or salt thereof, is in a buffer solution.